1 00:00:11,440 --> 00:00:13,640 Welcome to the Clinical Center Grand Rounds, 2 00:00:13,640 --> 00:00:17,440 a weekly series of educational lectures for physicians and 3 00:00:17,440 --> 00:00:20,080 health care professionals broadcast from the Clinical 4 00:00:20,080 --> 00:00:23,040 Center at the National Institutes of Health in 5 00:00:23,040 --> 00:00:24,840 Bethesda, MD. 6 00:00:24,840 --> 00:00:28,400 The NIH Clinical Center is the world's largest hospital totally 7 00:00:28,400 --> 00:00:32,080 dedicated to investigational research and leads the global 8 00:00:32,080 --> 00:00:35,040 effort in training today's investigators and discovering 9 00:00:35,040 --> 00:00:37,200 tomorrow's cures. 10 00:00:37,200 --> 00:00:47,040 Learn more by visiting us online at http://clinicalcenter.nih.gov 11 00:00:47,040 --> 00:00:48,880 I HAVE THE GREAT HONOR TO 12 00:00:48,880 --> 00:00:52,360 PRESENT OUR SPEAKER FOR TODAY, 13 00:00:52,360 --> 00:00:55,160 CHARLES E EGWUAGU, Ph.D. 14 00:00:55,160 --> 00:00:58,120 DR. EGWUAGU IS AN EPIDEMIOLOGIST 15 00:00:58,120 --> 00:01:00,160 AND IMMUNOLOGIST, AND THE CHIEF 16 00:01:00,160 --> 00:01:02,800 OF THE MOLECULAR IMMUNOLOGY 17 00:01:02,800 --> 00:01:04,400 SECTION, NATIONAL EYE INSTITUTE. 18 00:01:04,400 --> 00:01:07,640 HE RECEIVED HIS Ph.D. IN 19 00:01:07,640 --> 00:01:09,480 EPIDEMIOLOGY AND MICROBIOLOGY 20 00:01:09,480 --> 00:01:12,240 AND MASTER'S OF PHILOSOPHY IN 21 00:01:12,240 --> 00:01:14,880 MOLECULAR EPIDEMIOLOGY FROM YALE 22 00:01:14,880 --> 00:01:15,160 UNIVERSITY. 23 00:01:15,160 --> 00:01:18,840 HE ALSO COMPLETED A MASTER'S OF 24 00:01:18,840 --> 00:01:21,440 PUBLIC HEALTH FROM YALE SCHOOL 25 00:01:21,440 --> 00:01:22,840 OF MEDICINE. 26 00:01:22,840 --> 00:01:24,800 FROM 1987 TO 1990, DR. EGWUAGU 27 00:01:24,800 --> 00:01:27,760 WAS A STAFF FELLOW FOR THE 28 00:01:27,760 --> 00:01:29,440 SECTION ON EXPERIMENTAL 29 00:01:29,440 --> 00:01:31,400 IMMUNOLOGY AT NEI. 30 00:01:31,400 --> 00:01:33,240 HE THEN SERVED AS COMMISSIONED 31 00:01:33,240 --> 00:01:34,240 OFFICER OF THE UNITED STATES 32 00:01:34,240 --> 00:01:36,880 PUBLIC HEALTH AS MUCH AS, 33 00:01:36,880 --> 00:01:38,600 ATTAINING RANK OF CAPTAIN. 34 00:01:38,600 --> 00:01:40,400 CURRENTLY HE'S A SENIOR 35 00:01:40,400 --> 00:01:43,200 INVESTIGATOR AND CHIEF OF THE 36 00:01:43,200 --> 00:01:47,480 MOLECULAR IMMUNOLOGY SECTION. 37 00:01:47,480 --> 00:01:48,480 DR. EGWUAGU'S LABORATORY SEEKS 38 00:01:48,480 --> 00:01:49,960 AN INTEGRATED UNDERSTANDING OF 39 00:01:49,960 --> 00:02:00,520 MOLECULAR AND CELL MECHANISMS. 40 00:02:05,680 --> 00:02:07,000 PARTICULAR INTEREST ON THIS 41 00:02:07,000 --> 00:02:17,680 CYTOKINE IS -- IS ON CYTOSINE , 42 00:02:19,720 --> 00:02:22,680 TO DEVELOP REGULATORY B CELL 43 00:02:22,680 --> 00:02:25,080 IMMUNOTHERAPY FOR AUTOIMMUNE AND 44 00:02:25,080 --> 00:02:27,400 NEURODEGENERATIVE DISEASES. 45 00:02:27,400 --> 00:02:29,560 HE'S A RECIPIENT OF UNITED 46 00:02:29,560 --> 00:02:34,320 STATES PUBLIC HEALTH SERVICE 47 00:02:34,320 --> 00:02:38,680 COME COMMENDATION MEDAL AND 48 00:02:38,680 --> 00:02:42,160 DIRECTOR'S AWARD, AND MENTORING 49 00:02:42,160 --> 00:02:43,840 AWARD FOR HOWARD HUGHES MEDICAL 50 00:02:43,840 --> 00:02:45,240 INSTITUTE IN MONTGOMERY COUNTY 51 00:02:45,240 --> 00:02:46,000 SCHOOL SYSTEM AND NATIONAL 52 00:02:46,000 --> 00:02:48,520 INSTITUTES OF HEALTH INTERNSHIP 53 00:02:48,520 --> 00:02:48,760 PROGRAM. 54 00:02:48,760 --> 00:02:52,920 HE HAS OVER 100 PEER-REVIEWED 55 00:02:52,920 --> 00:02:54,640 PUBLICATIONS, SUPERVISED 60 56 00:02:54,640 --> 00:02:55,280 TRAINEES, FELLOWS, STUDENTS, A 57 00:02:55,280 --> 00:02:59,080 MEMBER OF THE U.S. PUBLIC HEALTH 58 00:02:59,080 --> 00:03:01,040 SERVICE PROFESSIONAL 59 00:03:01,040 --> 00:03:02,120 ASSOCIATION, ASSOCIATION FOR 60 00:03:02,120 --> 00:03:05,520 RESEARCH IN VISION AND 61 00:03:05,520 --> 00:03:06,120 OPHTHALMOLOGY, AND AMERICAN 62 00:03:06,120 --> 00:03:08,160 ASSOCIATION FOR ADVANCEMENT OF 63 00:03:08,160 --> 00:03:11,120 SCIENCE AND AMERICAN ASSOCIATION 64 00:03:11,120 --> 00:03:11,920 OF IMMUNOLOGISTS. 65 00:03:11,920 --> 00:03:19,480 HE IS A STANDING MEMBER OF STUY 66 00:03:19,480 --> 00:03:21,120 SECTION AND NIH CENTRAL TENURE 67 00:03:21,120 --> 00:03:21,680 COMMITTEE. 68 00:03:21,680 --> 00:03:24,240 THE TITLE OF THE PRESENTATION 69 00:03:24,240 --> 00:03:30,000 TODAY IS REGULATORY B CELLS AND 70 00:03:30,000 --> 00:03:31,880 EXOSOMES, UVEITIS, MULTIPLE 71 00:03:31,880 --> 00:03:32,000 SCLEROSIS AND GRAFT-VERSUS-HOST 72 00:03:32,000 --> 00:03:33,520 DISEASE. 73 00:03:33,520 --> 00:03:34,720 JOIN ME IN WELCOMING TODAY'S 74 00:03:34,720 --> 00:03:35,960 GRAND ROUNDS SPEAKER, DR. 75 00:03:35,960 --> 00:03:38,360 EGWUAGU. 76 00:03:38,360 --> 00:03:42,440 >> WELL, I HAVE NO FINANCIAL 77 00:03:42,440 --> 00:03:42,920 DISCLOSURES. 78 00:03:42,920 --> 00:03:46,200 AND I WOULD LIKE TO THANK YOU 79 00:03:46,200 --> 00:03:48,120 FOR THAT WONDERFUL INTRODUCTION. 80 00:03:48,120 --> 00:03:51,000 ALSO TO THANK THE NIH CLINICAL 81 00:03:51,000 --> 00:03:52,640 CENTER GRAND ROUNDS COMMITTEE 82 00:03:52,640 --> 00:03:56,520 FOR THE INVITATION TO SPEAK HERE 83 00:03:56,520 --> 00:03:59,840 THIS AFTERNOON. 84 00:03:59,840 --> 00:04:03,320 RESEARCH IN MY LAB OVER SEVERAL 85 00:04:03,320 --> 00:04:05,640 YEARS FOCUSED ON AUTOIMMUNE 86 00:04:05,640 --> 00:04:06,160 DISEASES. 87 00:04:06,160 --> 00:04:09,360 WE ARE PARTICULARLY INTERESTED 88 00:04:09,360 --> 00:04:13,760 IN UNDERSTANDING WHAT CAUSES AN 89 00:04:13,760 --> 00:04:16,240 ORGAN-SPECIFIC AUTOIMMUNE 90 00:04:16,240 --> 00:04:17,040 DISEASE SUCH AS UVEITIS OR 91 00:04:17,040 --> 00:04:18,120 MULTIPLE SCLEROSIS. 92 00:04:18,120 --> 00:04:21,600 OUR GOAL HAS BEEN TO IDENTIFY 93 00:04:21,600 --> 00:04:23,360 PATHOGENIC LYMPHOCYTES THAT FUEL 94 00:04:23,360 --> 00:04:27,000 THE VICIOUS CYCLES OF RELAPSING 95 00:04:27,000 --> 00:04:29,600 AND REMITTING INFLAMMATION THAT 96 00:04:29,600 --> 00:04:32,800 CHARACTERIZES UVEITIS AND 97 00:04:32,800 --> 00:04:33,520 MULTIPLE SCLEROSIS. 98 00:04:33,520 --> 00:04:38,160 THE ROLE OF T CELLS IN UVEITIS 99 00:04:38,160 --> 00:04:42,120 AND M.S. IS NOW WELL 100 00:04:42,120 --> 00:04:42,760 ESTABLISHED. 101 00:04:42,760 --> 00:04:45,480 WE PUBLISHED A PAPER THAT 102 00:04:45,480 --> 00:04:47,760 PROVIDED EVIDENCE FOR THE ROLE 103 00:04:47,760 --> 00:04:50,960 OF Th17 CELLS IN HUMAN 104 00:04:50,960 --> 00:04:51,280 UVEITIS. 105 00:04:51,280 --> 00:04:54,160 WE AND OTHERS HAVE ALSO 106 00:04:54,160 --> 00:04:57,400 IMPLICATED Th17 CELLS IN MOUSE 107 00:04:57,400 --> 00:05:01,680 MODELS OF MULTIPLE SCLEROSIS. 108 00:05:01,680 --> 00:05:02,440 HOWEVER, THIS AFTERNOON I WILL 109 00:05:02,440 --> 00:05:06,480 TALK ABOUT THE ROLE OF B CELLS 110 00:05:06,480 --> 00:05:09,800 IN SUPPRESSING CNS AUTOIMMUNE 111 00:05:09,800 --> 00:05:10,080 DISEASES. 112 00:05:10,080 --> 00:05:12,680 WITH PARTICULAR FOCUS ON OUR 113 00:05:12,680 --> 00:05:15,080 EFFORTS TO DEVELOP SAFE AND 114 00:05:15,080 --> 00:05:18,240 EFFECTIVE B CELL IMMUNOTHERAPY 115 00:05:18,240 --> 00:05:19,880 FOR UVEITIS. 116 00:05:19,880 --> 00:05:21,880 A GROUP OF POTENTIALLY BLINDING 117 00:05:21,880 --> 00:05:28,920 DISEASES WHICH CAN BE OF 118 00:05:28,920 --> 00:05:31,400 INFECTIOUS OR AUTOIMMUNE 119 00:05:31,400 --> 00:05:33,560 ETIOLOGY. 120 00:05:33,560 --> 00:05:39,480 UVEITIS IS A HETEROGENEOUS 121 00:05:39,480 --> 00:05:41,000 DISEASE DISCUSSING VKH, AND MANY 122 00:05:41,000 --> 00:05:42,640 OTHERS, CAN MANIFEST IN THE 123 00:05:42,640 --> 00:05:45,400 FRONT OF THE EYE AS ANTERIOR 124 00:05:45,400 --> 00:05:45,640 UVEITIS. 125 00:05:45,640 --> 00:05:52,720 IN THE BACK OF THE EYES, AS 126 00:05:52,720 --> 00:05:56,120 POSTERIOR UVEITIS. 127 00:05:56,120 --> 00:05:59,280 OR THROUGHOUT UVEA AS PAN 128 00:05:59,280 --> 00:06:02,920 UVEITIS. 129 00:06:02,920 --> 00:06:08,040 I'VE LISTED CURRENT TREATMENT. 130 00:06:08,040 --> 00:06:13,560 HOWEVER STEROIDS ARE STANDARD F 131 00:06:13,560 --> 00:06:14,040 CARE. 132 00:06:14,040 --> 00:06:21,840 THIS HAS BEEN THE IMPETUS TO 133 00:06:21,840 --> 00:06:23,920 DEVELOP EFFECTIVE NEW DRUGS FOR 134 00:06:23,920 --> 00:06:25,920 THESE POTENTIALLY BLINDING 135 00:06:25,920 --> 00:06:27,440 DISEASES. 136 00:06:27,440 --> 00:06:29,840 WE BELIEVE THAT BIOLOGICS AND 137 00:06:29,840 --> 00:06:32,440 CELL-BASED THERAPIES MAY BE 138 00:06:32,440 --> 00:06:35,000 SAFER ALTERNATIVES TO STEROIDS 139 00:06:35,000 --> 00:06:37,720 FOR TREATMENT OF UVEITIS. 140 00:06:37,720 --> 00:06:41,880 WE'RE THEREFORE FOCUSED ON 141 00:06:41,880 --> 00:06:44,400 MODULATING ACTIVITIES OF IL-12 142 00:06:44,400 --> 00:06:46,600 CYTOKINE FAMILY OF CYTOKINES, 143 00:06:46,600 --> 00:06:48,560 BECAUSE THEY PLAY CRITICAL ROLES 144 00:06:48,560 --> 00:06:49,760 IN HOST IMMUNITY. 145 00:06:49,760 --> 00:06:52,400 THIS INITIATE INNATE AS WELL AS 146 00:06:52,400 --> 00:06:53,360 ADAPTIVE IMMUNE RESPONSES. 147 00:06:53,360 --> 00:06:57,200 AND THEY ALSO REGULATE THE 148 00:06:57,200 --> 00:07:00,480 INTENSITY AND DURATION OF IMMUNE 149 00:07:00,480 --> 00:07:01,760 RESPONSES. 150 00:07:01,760 --> 00:07:04,840 THEY ALSO SKEW THE 151 00:07:04,840 --> 00:07:07,400 DIFFERENTIATION OF NAIVE 152 00:07:07,400 --> 00:07:09,200 LYMPHOCYTES TOWARDS PATHOGENIC 153 00:07:09,200 --> 00:07:10,640 OR IMMUNOSUPPRESSIVE SUBSETS OF 154 00:07:10,640 --> 00:07:11,000 LYMPHOCYTES. 155 00:07:11,000 --> 00:07:15,240 OUR GOAL HAS BEEN TO MITIGATE 156 00:07:15,240 --> 00:07:19,960 YOU'VE SITES BY TARGETING IL-12 157 00:07:19,960 --> 00:07:20,880 CYTOKINE SIGNALING PATHWAYS THAT 158 00:07:20,880 --> 00:07:24,880 WE CAN USE TO PROMOTE EXPANSION 159 00:07:24,880 --> 00:07:28,280 OF LYMPHOCYTES THAT SECRETE 160 00:07:28,280 --> 00:07:31,560 ANTI-INFLAMMATORY CYTOKINES. 161 00:07:31,560 --> 00:07:34,960 THE IL-12 FAMILY IS COMPRISED OF 162 00:07:34,960 --> 00:07:38,360 FOUR MEMBERS. 163 00:07:38,360 --> 00:07:42,040 IL-12 AND IL-23 ARE PRO INFLAM 164 00:07:42,040 --> 00:07:43,480 TEAR. 165 00:07:43,480 --> 00:07:44,920 IL-27 AND 37 ARE 166 00:07:44,920 --> 00:07:45,320 ANTI-INFLAMMATORY. 167 00:07:45,320 --> 00:07:49,400 EACH MEMBER OF THE IL-12 FAMILY 168 00:07:49,400 --> 00:07:52,680 CYTOKINE IS COMPOSED OF AN ALPHA 169 00:07:52,680 --> 00:07:55,640 CHAIN AND BETA SUBUNIT, BETA 170 00:07:55,640 --> 00:07:56,240 CHAIN. 171 00:07:56,240 --> 00:08:00,160 THE ALPHA SUBUNITS ARE P 35, 172 00:08:00,160 --> 00:08:03,640 P19, AND P 28. 173 00:08:03,640 --> 00:08:05,800 THESE ARE STRUCTURALLY SIMILAR 174 00:08:05,800 --> 00:08:09,760 TO IL-12 SUPER FAMILY CYTOKINES 175 00:08:09,760 --> 00:08:12,520 THAT INCLUDE ONCOSTATIC M, 176 00:08:12,520 --> 00:08:15,800 LEUKEMIA INHIBITORY FACTOR, IL-6 177 00:08:15,800 --> 00:08:19,200 AND MANY OTHERS. 178 00:08:19,200 --> 00:08:29,880 THE BETA SUBUNITS WITH H P 40, 179 00:08:29,880 --> 00:08:33,400 SIMILAR TO INTERFERON TYPE 1 180 00:08:33,400 --> 00:08:37,880 CYTOKINE RECEPTORS. 181 00:08:37,880 --> 00:08:41,520 THE PROFOUND INFLUENCE OF IL-12 182 00:08:41,520 --> 00:08:42,560 FAMILY CYTOKINES ON HOST 183 00:08:42,560 --> 00:08:44,640 IMMUNITY DERIVES FROM THE FACT 184 00:08:44,640 --> 00:08:49,240 THAT EACH OF THE ALPHA AND BETA 185 00:08:49,240 --> 00:08:52,680 SUBUNITS OF THE IL-12 FAMILY 186 00:08:52,680 --> 00:08:54,320 CYTOKINES, THE IMMEDIATE TARGETS 187 00:08:54,320 --> 00:08:56,840 OF TRANSCRIPTION FACTORS THAT 188 00:08:56,840 --> 00:08:58,800 ARE ACTIVATED BY DENDRITIC 189 00:08:58,800 --> 00:09:01,520 CELLS, DURING ANTIGEN PRIME IN 190 00:09:01,520 --> 00:09:03,840 THE LYMPH NODES. 191 00:09:03,840 --> 00:09:05,960 FOR EXAMPLE, PATHOGEN OF 192 00:09:05,960 --> 00:09:06,560 INTEREST IS A GRAM-NEGATIVE 193 00:09:06,560 --> 00:09:12,680 BACTERIA SUCH AS THE ONE THAT 194 00:09:12,680 --> 00:09:14,920 CAUSES MYCOBACTERIA, THAT CAUSES 195 00:09:14,920 --> 00:09:18,800 TUBERCULOSIS, THESE ARE GRAM 196 00:09:18,800 --> 00:09:20,480 POSITIVE BACTERIA, GRAM-NEGATIVE 197 00:09:20,480 --> 00:09:24,520 BACTERIA THAT HAPPEN ON CELL 198 00:09:24,520 --> 00:09:27,040 SURFACE, LIP OH POLYSACCHARIDES 199 00:09:27,040 --> 00:09:29,120 ENGAGE TLR 4 RECEPTORS ON THE 200 00:09:29,120 --> 00:09:30,440 ACTIVITY OF DENDRITIC CELLS 201 00:09:30,440 --> 00:09:32,400 CAUSING THEM TO INDUCE 202 00:09:32,400 --> 00:09:33,680 TRANSCRIPTION FACTOR THAT 203 00:09:33,680 --> 00:09:37,440 MOMENTS SECRETION OF IL-12 204 00:09:37,440 --> 00:09:37,800 CYTOKINES. 205 00:09:37,800 --> 00:09:39,720 THAT PROMOTES THE -- THAT 206 00:09:39,720 --> 00:09:50,280 PROMOTES THE DEVELOPMENT OF TH1 207 00:09:50,520 --> 00:09:51,200 SUBSETS. 208 00:09:51,200 --> 00:09:52,120 HOWEVER, IF THE PATHOGEN IS GRAM 209 00:09:52,120 --> 00:09:59,800 POSITIVE BACTERIA SUCH AS CAUSS 210 00:09:59,800 --> 00:10:01,560 STREP THROATS, THOSE BUGS 211 00:10:01,560 --> 00:10:05,840 CONTAIN GLYCAN ON CELL SURFACES, 212 00:10:05,840 --> 00:10:09,120 THAT INTERACTS CAN TLR2 213 00:10:09,120 --> 00:10:11,920 RECEPTORS IN THE CELL, INDUCING 214 00:10:11,920 --> 00:10:14,120 THEM TO ACTIVATE TRANSCRIPTION 215 00:10:14,120 --> 00:10:16,760 FACTORS THAT PROMOTE PRODUCTION 216 00:10:16,760 --> 00:10:21,160 OF HIGH LEVELS OF IL-23. 217 00:10:21,160 --> 00:10:27,520 IL-23 PROMOTES IMMUNE RESPONSE 218 00:10:27,520 --> 00:10:29,920 TOWARDS TH17 DEVELOPMENTAL 219 00:10:29,920 --> 00:10:32,560 PATHWAY. 220 00:10:32,560 --> 00:10:34,480 ON THAT CHRONIC INFLAMMATION 221 00:10:34,480 --> 00:10:45,040 DENDRITIC CELLS BEGIN TO PRODUCE 222 00:10:52,120 --> 00:10:52,800 IL-27. 223 00:10:52,800 --> 00:10:56,640 FOR THIS SEMINAR I WILL BEGIN BY 224 00:10:56,640 --> 00:10:59,280 PRESENTING OUR DATA, WE SHOW 225 00:10:59,280 --> 00:11:02,360 IL-27 AND IL-35 ARE AFFECTED 226 00:11:02,360 --> 00:11:03,240 BIOLOGICS. 227 00:11:03,240 --> 00:11:06,520 FOR TREATMENT OF UVEITIS OR 228 00:11:06,520 --> 00:11:07,400 ENCEPHALOMYELITIS. 229 00:11:07,400 --> 00:11:10,880 MOST OF OUR STUDIES ARE DONE IN 230 00:11:10,880 --> 00:11:11,080 MICE. 231 00:11:11,080 --> 00:11:13,520 I WOULD ALSO DISCUSS HOW STUDIES 232 00:11:13,520 --> 00:11:16,720 ON THE SUPPRESSION OF UVEITIS BY 233 00:11:16,720 --> 00:11:19,240 THESE CYTOKINES LED TO OUR 234 00:11:19,240 --> 00:11:26,400 DISCOVERY OF TWO NEW REGULATORY 235 00:11:26,400 --> 00:11:29,080 B CELL POPULATION, IN THE 236 00:11:29,080 --> 00:11:32,920 PERITONEAL CAVITY AND HUMAN 237 00:11:32,920 --> 00:11:34,440 UMBILICAL CORD. 238 00:11:34,440 --> 00:11:41,160 THAT SUPPRESSING INFLAMMATION BY 239 00:11:41,160 --> 00:11:48,920 SECRETING IL-27, I-27-BREGS. 240 00:11:48,920 --> 00:11:51,720 THESE CELLS RESIDE MOSTLY IN THE 241 00:11:51,720 --> 00:11:53,920 SPLEEN AND PERIPHERAL LYMPHOID 242 00:11:53,920 --> 00:11:54,120 TISSUES. 243 00:11:54,120 --> 00:11:57,560 THIS SUPPRESSED INFLAMMATION BY 244 00:11:57,560 --> 00:12:02,120 PRODUCING IL-35, AND WE CALL 245 00:12:02,120 --> 00:12:08,040 THESE B REGS I 35 B REGS. 246 00:12:08,040 --> 00:12:11,760 I'LL DISCUSS HOW STUDIES LED TO 247 00:12:11,760 --> 00:12:14,040 A DISCOVERY OF EXOSOMES THAT 248 00:12:14,040 --> 00:12:19,200 CONTAIN IL-27 AND WE'VE NAMED 249 00:12:19,200 --> 00:12:26,200 THEM I27 EXOSOMES, AND B REGS 250 00:12:26,200 --> 00:12:32,440 PRODUCE EXOSOMES THAT CONTAIN 251 00:12:32,440 --> 00:12:36,960 IL-35, I35-EXOSOMES, SUPPRESSING 252 00:12:36,960 --> 00:12:38,240 UVEITIS AND GRAFT-VERSUS-HOST 253 00:12:38,240 --> 00:12:40,880 DISEASE AS STAND-ALONE THERAPY 254 00:12:40,880 --> 00:12:47,560 OR IN COMBINATION WITH THE 255 00:12:47,560 --> 00:12:48,560 CORRESPONDING B REGS THAT 256 00:12:48,560 --> 00:12:50,200 PRODUCE THEM. 257 00:12:50,200 --> 00:12:51,280 I'LL CONCLUDE WITH UNPUBLISHED 258 00:12:51,280 --> 00:12:54,560 DATA PROVIDING NEW INSIGHT ON 259 00:12:54,560 --> 00:13:01,920 IMMUNOBIOLOGY AND BIOSYNTHESIS 260 00:13:01,920 --> 00:13:04,960 OF IL-27 AND IL-35. 261 00:13:04,960 --> 00:13:07,560 AFTER DISCOVERY OF IL-35, THERE 262 00:13:07,560 --> 00:13:11,120 WAS SIGNIFICANT -- HAD BEEN 263 00:13:11,120 --> 00:13:12,840 SIGNIFICANT INTEREST IN USING 264 00:13:12,840 --> 00:13:13,760 THEM THERAPEUTICALLY. 265 00:13:13,760 --> 00:13:15,240 AND THE FIRST ONE OF THE 266 00:13:15,240 --> 00:13:17,880 QUESTIONS WE ASKED WAS IF WE CAN 267 00:13:17,880 --> 00:13:22,280 USE IL-35 TO SUPPRESS UVEITIS. 268 00:13:22,280 --> 00:13:25,560 THE OBSTACLE TO ADDRESSING THIS 269 00:13:25,560 --> 00:13:30,680 ISSUE WAS THAT AT THAT TIME, 270 00:13:30,680 --> 00:13:32,680 BIOLOGIC -- BIOACTIVE IL-35 WAS 271 00:13:32,680 --> 00:13:36,080 NOT AVAILABLE COMMERCIALLY SO WE 272 00:13:36,080 --> 00:13:38,920 HAD TO GENETICALLY ENGINEER 273 00:13:38,920 --> 00:13:40,120 RECOMBINANT IL-35 FOR THESE 274 00:13:40,120 --> 00:13:40,360 STUDIES. 275 00:13:40,360 --> 00:13:49,720 WHAT I'M SHOWING YOU HERE IS 276 00:13:49,720 --> 00:13:51,480 CONSTRUCT THAT WE PRODUCED AND 277 00:13:51,480 --> 00:13:56,720 USED TO EXPRESS THE IL-35 IN 278 00:13:56,720 --> 00:14:00,120 CELLS. 279 00:14:00,120 --> 00:14:04,280 CONSTRUCT CONSISTS OF P 35 280 00:14:04,280 --> 00:14:08,440 cDNA, BOTH JOINED BY IRES, 281 00:14:08,440 --> 00:14:10,920 WHICH IS AN RNA ELEMENT THAT 282 00:14:10,920 --> 00:14:13,720 ALLOWS US TO EXPRESS THIS 283 00:14:13,720 --> 00:14:15,360 PROTEIN AND SECRETE 284 00:14:15,360 --> 00:14:16,200 INDEPENDENTLY. 285 00:14:16,200 --> 00:14:19,280 SO, WE CLONED THIS CONSTRUCT 286 00:14:19,280 --> 00:14:22,120 INTO THE PMI EFFECTOR USE TO 287 00:14:22,120 --> 00:14:24,320 TRANSFECT THESE CELLS TO 288 00:14:24,320 --> 00:14:25,720 GENERATE RECOMBINANT PROTEIN. 289 00:14:25,720 --> 00:14:27,920 THE PURPOSE OF THIS SLIDE HERE 290 00:14:27,920 --> 00:14:33,080 IS JUST TO SHOW YOU THAT WE 291 00:14:33,080 --> 00:14:37,000 PRODUCED HIGHLY PURIFIED IL-35 292 00:14:37,000 --> 00:14:38,440 CYTOKINE. 293 00:14:38,440 --> 00:14:48,520 AND WE DEMONSTRATE HERE THAT THE 294 00:14:48,520 --> 00:14:54,960 IL-35 IS HETERODIMERIC CYTOKINE 295 00:14:54,960 --> 00:15:01,320 BY SEDIMENTATION, EQUILIBRIUM OR 296 00:15:01,320 --> 00:15:02,720 CENTRIFUGATION. 297 00:15:02,720 --> 00:15:06,120 THEY MIGRATE AS HETERODIMER OF 298 00:15:06,120 --> 00:15:08,440 57.7 KILODALTON. 299 00:15:08,440 --> 00:15:11,920 ON THAT REDUCED CONDITION, THEY 300 00:15:11,920 --> 00:15:14,560 MIGRATE AS INDIVIDUAL SUBUNITS. 301 00:15:14,560 --> 00:15:19,360 AND THIS WAS ONE OF THE FIRST 302 00:15:19,360 --> 00:15:20,440 PRODUCTION OF BIOACTIVE 303 00:15:20,440 --> 00:15:21,040 HETERODIMERIC CYTOKINES THAT 304 00:15:21,040 --> 00:15:23,120 COULD BE USED FOR THESE SORT OF 305 00:15:23,120 --> 00:15:24,600 STUDIES. 306 00:15:24,600 --> 00:15:28,120 SO THE FIRST THING WE DID WAS TO 307 00:15:28,120 --> 00:15:30,880 ESTABLISH THE IL-35 IS INDEED 308 00:15:30,880 --> 00:15:31,720 BIOLOGICALLY ACTIVE. 309 00:15:31,720 --> 00:15:34,480 AND WE DID SO BY SHOWING THAT 310 00:15:34,480 --> 00:15:38,760 THE IL-35 CAN SUPPRESS PRIMARY B 311 00:15:38,760 --> 00:15:40,960 CELLS PROLIFERATION, AND DID SO 312 00:15:40,960 --> 00:15:43,800 BY INDUCING B REGS TO PRODUCE 313 00:15:43,800 --> 00:15:47,960 HIGH LEVELS OF IL-10, WHICH IS 314 00:15:47,960 --> 00:15:51,320 AN IMMUNE SUPPRESSIVE CYTOKINE. 315 00:15:51,320 --> 00:15:54,600 INTERESTINGLY, WHY THE IL-35 CAN 316 00:15:54,600 --> 00:15:57,360 SUPPRESS CONVERSIONAL B CELLS, 317 00:15:57,360 --> 00:15:59,560 CAN SUPPRESS PROLIFERATION, IT 318 00:15:59,560 --> 00:16:02,000 INDUCES THE EXPANSION OF VERY 319 00:16:02,000 --> 00:16:05,600 RARE B CELL POPULATION, THAT 320 00:16:05,600 --> 00:16:06,880 PRODUCES IMMUNOSUPPRESSIVE 321 00:16:06,880 --> 00:16:07,760 CYTOKINE IL-10. 322 00:16:07,760 --> 00:16:14,240 AND WE CALL THIS IL-10 B-REGS. 323 00:16:14,240 --> 00:16:16,080 THE LOGICAL EXTENSION OF THIS 324 00:16:16,080 --> 00:16:21,120 WORK IS TO FIND OUT WHETHER WE 325 00:16:21,120 --> 00:16:22,760 COULD USE IL-35 TO SUPPRESS 326 00:16:22,760 --> 00:16:24,960 INFLAMMATIONS IN VIVO AND FOR 327 00:16:24,960 --> 00:16:29,280 THIS WE USED THE EXPERIMENTAL 328 00:16:29,280 --> 00:16:30,440 AUTOIMMUNE UVEITIS MODEL. 329 00:16:30,440 --> 00:16:36,480 EAU IS THE MODEL OF HUMAN 330 00:16:36,480 --> 00:16:37,800 UVEITIS, SHARES 331 00:16:37,800 --> 00:16:48,480 HISTOIMMUNOLOGYIC FEATURES WITH 332 00:16:48,480 --> 00:16:51,040 HUMAN DISEASE, INDUCES MOUSE 333 00:16:51,040 --> 00:16:55,000 STRAINS. 334 00:16:55,000 --> 00:16:58,840 SYMPTOMS OF EAU APPEARS 8, 12, 335 00:16:58,840 --> 00:17:02,240 TO 14 POST IMMUNIZATION, AND 336 00:17:02,240 --> 00:17:05,840 TARGETED DESTRUCTION OF THE 337 00:17:05,840 --> 00:17:09,440 PHOTORECEPTOR LAYER. 338 00:17:09,440 --> 00:17:12,000 IN THE MOUSE WE SEE UNDULATING 339 00:17:12,000 --> 00:17:17,600 CELLS IN THE PHOTORESETTIVE 340 00:17:17,600 --> 00:17:21,880 LAYER, A HALLMARK FEATURE OF 341 00:17:21,880 --> 00:17:24,880 SEVERE UVEITIS IN MICE. 342 00:17:24,880 --> 00:17:31,800 IN THIS STUDY, WE INDUCE EAU IN 343 00:17:31,800 --> 00:17:33,560 MICE, AT THE SAME TIME STARTED 344 00:17:33,560 --> 00:17:36,960 TREATING THEM THE SAME TIME WE 345 00:17:36,960 --> 00:17:40,000 INDUCED EAU STARTED TREATING 346 00:17:40,000 --> 00:17:44,080 THEM WITH 100 NANOGRAMS OF THE 347 00:17:44,080 --> 00:17:46,600 RECOMBINANT IL-35, OR ALSO GAVE 348 00:17:46,600 --> 00:17:48,360 THEM 100 NANOGRAMS OF PMIB, 349 00:17:48,360 --> 00:17:51,720 WHICH IS A CONTROL. 350 00:17:51,720 --> 00:17:57,520 PMIB IS AN EXTRACT DERIVED FROM 351 00:17:57,520 --> 00:18:08,000 CELLS TRANSFECTED WITH MTPMI 352 00:18:13,600 --> 00:18:18,880 DEFECTOR, BY FUNDUSCOPY, 353 00:18:18,880 --> 00:18:20,400 HISTOLOGY, CYTOKINE STAINING, 354 00:18:20,400 --> 00:18:20,640 ANALYSIS. 355 00:18:20,640 --> 00:18:22,480 WHEN YOU LOOK AT FUNDUS IMAGE, 356 00:18:22,480 --> 00:18:24,680 IS THIS WHAT THE FORMAL FUNDUS 357 00:18:24,680 --> 00:18:28,840 LOOKS LIKE, IT HAS THIS 358 00:18:28,840 --> 00:18:31,000 WELL-DEFINED OPTIC DISC. 359 00:18:31,000 --> 00:18:34,320 DURING INFLAMMATION OR IN EAU 360 00:18:34,320 --> 00:18:39,000 THERE'S A SWELLING OF THE OPTIC 361 00:18:39,000 --> 00:18:43,040 DISC, AND WHICH WE CALL PAPULAR 362 00:18:43,040 --> 00:18:43,400 EDEMA. 363 00:18:43,400 --> 00:18:45,480 THESE MICE SHOWN HERE IN THIS 364 00:18:45,480 --> 00:18:50,400 FUNDUS IMAGE IN BLUE, THEY ALSO 365 00:18:50,400 --> 00:18:52,720 DEVELOP RETINAL VASCULITIS, AND 366 00:18:52,720 --> 00:18:58,880 THE WHITE ARROW HERE SHOWS AREAS 367 00:18:58,880 --> 00:19:01,040 OF RETINAL AND CORONAL 368 00:19:01,040 --> 00:19:03,120 INFILTRATES, HALLMARK FEATURES 369 00:19:03,120 --> 00:19:06,160 OF UVEITIS. 370 00:19:06,160 --> 00:19:08,920 AND IN THE HISTOLOGY, STAINING 371 00:19:08,920 --> 00:19:11,960 HERE, THESE AREAS MARKED IN BLUE 372 00:19:11,960 --> 00:19:15,720 ASTERISKS SHOW AREAS OF RETINAL 373 00:19:15,720 --> 00:19:17,560 INFOLDING, WHICH I MENTIONED 374 00:19:17,560 --> 00:19:19,160 EARLIER IS HALLMARK FEATURE OF 375 00:19:19,160 --> 00:19:21,400 SEVERE UVEITIS IN THE MOUSE. 376 00:19:21,400 --> 00:19:24,000 NOW, WHEN YOU LOOK AT THE MICE 377 00:19:24,000 --> 00:19:28,920 THAT WERE TREATED WITH IL-35, 378 00:19:28,920 --> 00:19:31,040 THEY WERE PROTECTED FROM THESE 379 00:19:31,040 --> 00:19:34,080 CARDINAL FEATURES OF UVEITIS 380 00:19:34,080 --> 00:19:34,840 I'VE JUST SHOWN YOU. 381 00:19:34,840 --> 00:19:37,880 WE BELIEVE WE FOUND OUT THAT THE 382 00:19:37,880 --> 00:19:39,320 SUPPRESSION OF UVEITIS DERIVED 383 00:19:39,320 --> 00:19:43,800 FROM THE FACTS THAT IL-35 384 00:19:43,800 --> 00:19:47,040 SUPPRESSED PATHOGENIC CELLS THAT 385 00:19:47,040 --> 00:19:48,400 PRODUCED THE PATHOGENIC 386 00:19:48,400 --> 00:19:50,280 LYMPHOCYTES IL-17 AND INTERFERON 387 00:19:50,280 --> 00:19:50,600 GAMMA. 388 00:19:50,600 --> 00:19:55,440 AT THE SAME TIME THE IL-35 389 00:19:55,440 --> 00:19:59,680 INDUCED EXPANSION OF LYMPHOCYTES 390 00:19:59,680 --> 00:20:00,920 THAT PRODUCED IL-10. 391 00:20:00,920 --> 00:20:01,880 SURPRISINGLY WHAT WE FOUND IS 392 00:20:01,880 --> 00:20:07,480 THAT MOST OF THE IL-10 PRODUCING 393 00:20:07,480 --> 00:20:12,320 B REGS ALSO PRODUCED IL-35, THE 394 00:20:12,320 --> 00:20:13,960 ONES FOUND IN LYMPH NODES 395 00:20:13,960 --> 00:20:19,320 PRODUCED EYE LEVELS OF IL-35. 396 00:20:19,320 --> 00:20:22,160 AS MUCH AS 64% ALSO PRODUCED 397 00:20:22,160 --> 00:20:22,480 IL-35. 398 00:20:22,480 --> 00:20:29,600 THESE ARE THE ONES THAT WE CALL 399 00:20:29,600 --> 00:20:31,440 I35 B-REGS. 400 00:20:31,440 --> 00:20:36,920 IN THIS STUDY, WE DECIDED TO 401 00:20:36,920 --> 00:20:39,440 ISOLATE THOSE B REGS TO EXAMINE 402 00:20:39,440 --> 00:20:43,400 WHETHER THEY CAN BE USED TO 403 00:20:43,400 --> 00:20:44,680 SUPPRESS AND ESTABLISH UVEITIS. 404 00:20:44,680 --> 00:20:46,400 WE ISOLATED CELLS FROM THE 405 00:20:46,400 --> 00:20:48,440 SPLEEN AND LYMPH NODES OF THE 406 00:20:48,440 --> 00:20:52,480 MICE THAT WERE TREATED WITH 407 00:20:52,480 --> 00:20:53,240 IL-35. 408 00:20:53,240 --> 00:20:56,960 AND THEN WE CULTURED THEM, 409 00:20:56,960 --> 00:21:06,040 REACTIVATED THEM WITH IRPP AND 410 00:21:06,040 --> 00:21:08,040 CD40, IL-10 CAPTURE REAGENT TO 411 00:21:08,040 --> 00:21:10,200 SORT THESE CELLS THAT MAKE IL-10 412 00:21:10,200 --> 00:21:13,280 AND ENRICH THEM, WE CAN ENRICH 413 00:21:13,280 --> 00:21:15,680 THEM AND SOME PREPARATION WOULD 414 00:21:15,680 --> 00:21:19,880 GET MORE THAN 90% OF 415 00:21:19,880 --> 00:21:21,040 IL-10-PRODUCING B REG CELLS. 416 00:21:21,040 --> 00:21:25,320 SO, WHAT WE DID IN THIS STUDY 417 00:21:25,320 --> 00:21:29,680 NOW IS TO INDUCE EAU AND GIVE IT 418 00:21:29,680 --> 00:21:31,880 FOUR DAYS LEAD PERIOD TO BUILD 419 00:21:31,880 --> 00:21:35,080 UP CRITICAL MASS OF THE 420 00:21:35,080 --> 00:21:36,160 POTENTIAL PATHOGENIC LYMPHOCYTES 421 00:21:36,160 --> 00:21:40,360 AND THEN HIT THEM WITH THE 10 422 00:21:40,360 --> 00:21:43,400 MILLION B REG CELLS. 423 00:21:43,400 --> 00:21:46,160 WE ALSO IMMUNIZED THEM WITH -- 424 00:21:46,160 --> 00:21:51,160 INJECTED THEM 10 MILLION B 425 00:21:51,160 --> 00:21:54,120 CELLS, IL-10, SOME MICE LEFT 426 00:21:54,120 --> 00:21:56,400 THEM UNTREATED WITH ANYTHING. 427 00:21:56,400 --> 00:21:59,160 NOW, WHEN YOU LOOK AT THE FUNDUS 428 00:21:59,160 --> 00:22:02,200 IMAGES, YOU SEE HERE THE 429 00:22:02,200 --> 00:22:03,520 RECAPITULATE ESSENTIAL FEATURES 430 00:22:03,520 --> 00:22:04,720 OF UVEITIS. 431 00:22:04,720 --> 00:22:07,800 LOOKING AT THE HISTOLOGY, IT 432 00:22:07,800 --> 00:22:11,400 GIVES YOU A MUCH MORE COMPLETE 433 00:22:11,400 --> 00:22:14,040 PICTURE. 434 00:22:14,040 --> 00:22:15,360 YOU HAVE SIGNIFICANT 435 00:22:15,360 --> 00:22:16,560 INFILTRATION OF INFLAMMATORY 436 00:22:16,560 --> 00:22:20,000 CELLS IN THE RETINA, IN THE 437 00:22:20,000 --> 00:22:23,680 IRIS, AND IN THE VITREOUS. 438 00:22:23,680 --> 00:22:27,880 AGAIN, YOU SEE THE CARDINAL 439 00:22:27,880 --> 00:22:29,800 FEATURES OF UNDULATING CELLS 440 00:22:29,800 --> 00:22:32,400 WHICH WE CALL RETINAL INFOLDING. 441 00:22:32,400 --> 00:22:36,360 WHAT THIS DATA SUGGESTS IS THAT 442 00:22:36,360 --> 00:22:38,560 B-REGS, THE ONES THAT PRODUCE EX 443 00:22:38,560 --> 00:22:46,320 VIVO, CAN BE USED TO SUPPRESS AN 444 00:22:46,320 --> 00:22:48,720 ESTABLISHED UVEITIS. 445 00:22:48,720 --> 00:22:50,040 WELL, IL-27 IS THE OTHER 446 00:22:50,040 --> 00:22:52,000 IMMUNOSUPPRESSIVE MEMBER OF THE 447 00:22:52,000 --> 00:22:53,800 IL-12 FAMILY. 448 00:22:53,800 --> 00:22:56,280 WE ALSO ASKED WHETHER IT CAN BE 449 00:22:56,280 --> 00:23:02,480 USED AS A BIOLOGIC FOR UVEITIS. 450 00:23:02,480 --> 00:23:05,960 OUR INTEREST IN IL-27 AS A 451 00:23:05,960 --> 00:23:08,360 BIOLOGIC FOR UVEITIS DERIVED 452 00:23:08,360 --> 00:23:10,880 FROM OUR PREVIOUS PUBLICATIONS, 453 00:23:10,880 --> 00:23:12,760 SOME SHOWN HERE, WHETHER WE 454 00:23:12,760 --> 00:23:15,400 SHOWED SUPPRESSION OF UVEITIS IN 455 00:23:15,400 --> 00:23:16,320 MICE CORRELATES WITH 456 00:23:16,320 --> 00:23:19,760 UPREGULATION OF IL-27 IN THE 457 00:23:19,760 --> 00:23:23,680 RETINA AND THIS BY MICROGLIAL 458 00:23:23,680 --> 00:23:24,440 CELLS. 459 00:23:24,440 --> 00:23:30,120 THE FACT THAT IL-27 AND IL-35 460 00:23:30,120 --> 00:23:30,600 ARE IMMUNE SUPPRESSIVE 461 00:23:30,600 --> 00:23:32,520 CYTOKINES, THEY ARE STRUCTURAL 462 00:23:32,520 --> 00:23:35,920 AND FUNCTIONALLY SIMILAR. 463 00:23:35,920 --> 00:23:39,400 WE HAVE HYPOTHESIZED IL-27 MAY 464 00:23:39,400 --> 00:23:41,080 ALSO BE AN EFFECTIVE BIOLOGIC 465 00:23:41,080 --> 00:23:45,360 FOR TREATMENT OF UVEITIS. 466 00:23:45,360 --> 00:23:50,160 WE ALSO POSITED THAT IL-27 467 00:23:50,160 --> 00:23:53,320 PRODUCING B REGS, IL-27 B REGS 468 00:23:53,320 --> 00:23:58,360 MUST ALSO EXIST IN VIVO AS I 35 469 00:23:58,360 --> 00:24:02,680 B REGS. 470 00:24:02,680 --> 00:24:08,040 IN THIS STUDY, WHAT WE DID WAS 471 00:24:08,040 --> 00:24:10,120 TO INDUCE EAU AND EXAMINE 472 00:24:10,120 --> 00:24:15,240 WHETHER IL-35 CAN BE -- CAN 473 00:24:15,240 --> 00:24:19,840 SERVE AS EFFECTIVE BIOLOGIC FOR 474 00:24:19,840 --> 00:24:21,480 MOUSE UVEITIS. 475 00:24:21,480 --> 00:24:23,480 MICE -- EAU WAS INDUCED, AT THE 476 00:24:23,480 --> 00:24:29,280 SAME TIME EAU WAS INDUCED, WE 477 00:24:29,280 --> 00:24:32,680 GAVE THEM IL-27, AT THE TIME OF 478 00:24:32,680 --> 00:24:33,640 EAU INDUCTION. 479 00:24:33,640 --> 00:24:37,240 AND EVERY OTHER DAY TILL DAY 10. 480 00:24:37,240 --> 00:24:39,440 AND WHAT IS SHOWN HERE IN THIS 481 00:24:39,440 --> 00:24:46,200 FUNDUS IMAGE IS THAT INDEED THE 482 00:24:46,200 --> 00:24:48,200 IL-27 CONVERTS PROTECTION 483 00:24:48,200 --> 00:24:53,000 AGAINST SEVERE UVEITIS SHOWN BY 484 00:24:53,000 --> 00:24:55,640 DECREASED DISEASE SCORE, 485 00:24:55,640 --> 00:24:58,440 RECAPITULATED BY HISTOLOGY SHOWN 486 00:24:58,440 --> 00:25:02,200 SIGNIFICANT INFLAMMATIONS IN THE 487 00:25:02,200 --> 00:25:05,600 PBS-TREATED MICE COMPARED TO 488 00:25:05,600 --> 00:25:07,680 IL-TREATED MICE. 489 00:25:07,680 --> 00:25:09,440 WE USE TOMOGRAPHY TO MONITOR EAU 490 00:25:09,440 --> 00:25:12,040 IN MICE. 491 00:25:12,040 --> 00:25:19,280 IT'S A NON-INVASIVE TECHNIQUE 492 00:25:19,280 --> 00:25:21,760 JUST LIKE FUNDUSCOPY, WITH 493 00:25:21,760 --> 00:25:28,240 RECRUITMENT OF INFLAMMATORY 494 00:25:28,240 --> 00:25:29,800 CELLS INTO THE RETINA. 495 00:25:29,800 --> 00:25:31,600 WE ADDRESSED NEXT QUESTION, 496 00:25:31,600 --> 00:25:34,680 WHETHER IL-27 B REGS DO INDEED 497 00:25:34,680 --> 00:25:35,800 EXIST IN VIVO. 498 00:25:35,800 --> 00:25:37,800 BEFORE ADDRESSING THIS ISSUE I 499 00:25:37,800 --> 00:25:39,760 WOULD LIKE TO MAKE -- TO NOTE 500 00:25:39,760 --> 00:25:45,120 THAT THERE ARE TWO TYPES OF B 501 00:25:45,120 --> 00:25:50,720 CELLS IN MAMMALS. 502 00:25:50,720 --> 00:25:56,520 ONE IS B-1 LINEAGE, AND B-2 503 00:25:56,520 --> 00:25:56,760 LINEAGE. 504 00:25:56,760 --> 00:25:59,000 AND B-2 CELLS, THEY DEVELOP IN 505 00:25:59,000 --> 00:26:01,240 THE BONE MARROW, THEY ARE THE 506 00:26:01,240 --> 00:26:03,400 MOST PLENTIFUL B CELLS THAT YOU 507 00:26:03,400 --> 00:26:06,360 FIND IN THE SPLEEN AND IN 508 00:26:06,360 --> 00:26:09,000 PERIPHERAL LYMPHOID TISSUES. 509 00:26:09,000 --> 00:26:12,800 AND THE MOST ALL OF IT, I 35 B 510 00:26:12,800 --> 00:26:17,000 REGS OR B REGS PRODUCING IL-35 511 00:26:17,000 --> 00:26:21,040 DISCUSSED EARLIER, ALL DERIVE 512 00:26:21,040 --> 00:26:22,760 FROM THE B-2 LINEAGE. 513 00:26:22,760 --> 00:26:25,720 WE WERE THEREFORE SURPRISED THAT 514 00:26:25,720 --> 00:26:29,240 THE IL-27 WAS NOT -- THESE B 515 00:26:29,240 --> 00:26:31,840 CELLS DO NOT MAKE IL-27. 516 00:26:31,840 --> 00:26:35,160 INSTEAD, WE FOUND THAT THE IL-27 517 00:26:35,160 --> 00:26:38,280 PRODUCTION WAS RESTRICTED TO THE 518 00:26:38,280 --> 00:26:42,240 B-1 LINEAGE, AND THIS B-1 CELLS, 519 00:26:42,240 --> 00:26:45,320 AND B-1 CELLS AT THE EARLIEST 520 00:26:45,320 --> 00:26:50,760 CELLS, B CELLS, THAT APPEAR IN 521 00:26:50,760 --> 00:26:53,840 DEVELOPMENT. 522 00:26:53,840 --> 00:26:55,600 DERIVED FROM EMBRYONIC YOLK SAC, 523 00:26:55,600 --> 00:26:57,920 RESIDING IN THE PERITONEAL 524 00:26:57,920 --> 00:27:00,200 CAVITY MOSTLY AND UMBILICAL CORD 525 00:27:00,200 --> 00:27:04,160 BLOOD OF HUMANS AND MICE. 526 00:27:04,160 --> 00:27:07,320 WE USE COMBINATION OF 527 00:27:07,320 --> 00:27:08,480 IMMUNOHISTOCHEMISTRY AND 528 00:27:08,480 --> 00:27:12,680 CONFOCAL MICROSCOPY TO 529 00:27:12,680 --> 00:27:15,520 DEMONSTRATE INDEED THE B-1A 530 00:27:15,520 --> 00:27:17,720 CELLS WHEN ACTIVATED, PRODUCE 531 00:27:17,720 --> 00:27:20,880 SIGNIFICANT AMOUNT OF IL-27, AS 532 00:27:20,880 --> 00:27:24,840 SHOWN HERE IN THIS CONFOCAL 533 00:27:24,840 --> 00:27:30,520 IMAGE HERE, AND SHOWN HERE WE 534 00:27:30,520 --> 00:27:35,240 DEMONSTRATE THAT THESE IL-27 MAY 535 00:27:35,240 --> 00:27:40,960 INDEED BE HETERODIMERIC ACTIVE 536 00:27:40,960 --> 00:27:41,600 BIOLOGICAL IL-27. 537 00:27:41,600 --> 00:27:44,400 BUT DEFINITELY PROOF CAME FROM 538 00:27:44,400 --> 00:27:46,200 PROXIMITY LIGATION ASSAY, THIS 539 00:27:46,200 --> 00:27:48,840 IS A VERY, VERY SENSITIVE 540 00:27:48,840 --> 00:27:51,760 TECHNIQUE THAT YOU USE TO DETECT 541 00:27:51,760 --> 00:27:55,200 TWO MOLECULES THAT ARE WITHIN 542 00:27:55,200 --> 00:27:59,120 EACH OTHER, WITHIN 40-NANOMETER 543 00:27:59,120 --> 00:28:01,200 FROM EACH OTHER IN THE MEMBRANE. 544 00:28:01,200 --> 00:28:03,800 WHAT YOU SEE HER IS THIS THESE 545 00:28:03,800 --> 00:28:08,520 RED DOTS, SHOWS THE AREA WHERE 546 00:28:08,520 --> 00:28:13,560 THE 28 AND 3 ARE IN CLOSE 547 00:28:13,560 --> 00:28:16,200 PROXIMITY TO EACH OTHER, LESS 548 00:28:16,200 --> 00:28:19,040 THAN 40 FROM EACH OTHER. 549 00:28:19,040 --> 00:28:22,440 WE ALSO IDENTIFIED THE IL-25, 550 00:28:22,440 --> 00:28:24,720 DETECTED THEM ALSO IN HUMAN 551 00:28:24,720 --> 00:28:28,800 BLOOD, BUT IN MOST ABUNDANCE 552 00:28:28,800 --> 00:28:36,560 AMOUNT OF IL-27 WAS FOUND IN THE 553 00:28:36,560 --> 00:28:38,720 UMBILICAL CORD BLOOD. 554 00:28:38,720 --> 00:28:41,160 WELL, IN THIS STUDY WHAT WE DID 555 00:28:41,160 --> 00:28:44,120 WAS REPEATED THE SAME SORT OF 556 00:28:44,120 --> 00:28:51,320 EXPERIMENT TO ASK HERE NOW 557 00:28:51,320 --> 00:28:52,640 WHETHER GENERATED B REGS CAN BE 558 00:28:52,640 --> 00:28:55,440 USED TO TREAT UVEITIS. 559 00:28:55,440 --> 00:28:59,240 THE ANSWER IS YES. 560 00:28:59,240 --> 00:29:05,360 WE OBTAINED -- GENERATED THE 561 00:29:05,360 --> 00:29:08,720 I27-BREGULARS AND SORT -- B-REGS 562 00:29:08,720 --> 00:29:09,720 AND SORTED THEM. 563 00:29:09,720 --> 00:29:13,960 THESE MICE HERE FROM THE FUNDUS 564 00:29:13,960 --> 00:29:16,080 IMAGES, THEY ARE PROTECTED FROM 565 00:29:16,080 --> 00:29:19,920 UVEITIS WHEN TREATED WITH THE 566 00:29:19,920 --> 00:29:24,400 I27-BREGS INDICATED BY LOWER EAU 567 00:29:24,400 --> 00:29:24,800 SCORES. 568 00:29:24,800 --> 00:29:27,040 AND WE FOUND THE SUPPRESSION OF 569 00:29:27,040 --> 00:29:31,720 THE DISEASE WAS NOT ONLY BY 570 00:29:31,720 --> 00:29:32,960 SUPPRESSING PATHOGENIC 571 00:29:32,960 --> 00:29:33,280 LYMPHOCYTES. 572 00:29:33,280 --> 00:29:36,800 THEY ALSO INDUCED PRODUCTION OF 573 00:29:36,800 --> 00:29:41,160 HIGH LEVELS OF CYTOKINES LIKE 574 00:29:41,160 --> 00:29:46,080 IL-10, IL-35, IL-27. 575 00:29:46,080 --> 00:29:47,960 AND THIS IS REMINISCENT OF WHAT 576 00:29:47,960 --> 00:29:50,720 IS REFERRED TO -- THAT I WOULD 577 00:29:50,720 --> 00:29:54,640 LIKE TO CALL INFECTIOUS 578 00:29:54,640 --> 00:29:54,960 TOLERANCE. 579 00:29:54,960 --> 00:29:56,920 HOWEVER, BECAUSE WE HAD A 580 00:29:56,920 --> 00:30:00,440 PRODUCTION OF IL-10 AND IL-35, 581 00:30:00,440 --> 00:30:04,840 TWO OF OUR REVIEWERS OF OUR 582 00:30:04,840 --> 00:30:06,800 PAPERS INSISTED WE PROVIDE 583 00:30:06,800 --> 00:30:08,880 EVIDENCE THIS DISEASE 584 00:30:08,880 --> 00:30:10,280 SUPPRESSION HERE IS NOT CAUSED 585 00:30:10,280 --> 00:30:11,480 BY THESE OTHER CYTOKINES. 586 00:30:11,480 --> 00:30:15,640 AND FOR THIS WE DECIDED TO USE A 587 00:30:15,640 --> 00:30:18,600 CRISPR TECHNOLOGY TO GENETICALLY 588 00:30:18,600 --> 00:30:23,000 ENGINEER MICE WITH TARGETED 589 00:30:23,000 --> 00:30:24,360 DELETION OF THE IL-27 RECEPTOR 590 00:30:24,360 --> 00:30:26,080 FROM THE B CELLS. 591 00:30:26,080 --> 00:30:30,920 NOW, WHEN WE REPEATED THIS 592 00:30:30,920 --> 00:30:33,000 EXPERIMENT WE FOUND THE BREGS 593 00:30:33,000 --> 00:30:37,560 CAN NO LONGER SUPPRESS DISEASE. 594 00:30:37,560 --> 00:30:40,560 SUGGESTING IN ADDITION TO ONE OF 595 00:30:40,560 --> 00:30:45,600 THE MECHANISMS USED IS BY 596 00:30:45,600 --> 00:30:46,920 INDUCING INFECTIOUS TOLERANCE 597 00:30:46,920 --> 00:30:55,000 WHICH IS A MECHANISM BY WHICH 598 00:30:55,000 --> 00:30:55,960 INFLAMMATORY CYTOKINES CAN 599 00:30:55,960 --> 00:30:58,840 CONVERT B CELLS TO REGULATORY 600 00:30:58,840 --> 00:31:00,400 CELLS, AND THAT'S WHAT WE'RE 601 00:31:00,400 --> 00:31:02,600 SEEING WITH INCREASE IN CELLS 602 00:31:02,600 --> 00:31:06,480 THAT COULD MAKE OTHER CYTOKINES, 603 00:31:06,480 --> 00:31:09,360 ALSO CAUSES EXPANSION OF IL-27. 604 00:31:09,360 --> 00:31:11,760 AND THIS IS WHAT'S RESPONSIBLE 605 00:31:11,760 --> 00:31:16,680 FOR THE SUPPRESSION OF THIS 606 00:31:16,680 --> 00:31:16,920 DISEASE. 607 00:31:16,920 --> 00:31:19,760 AT THE BEGINNING I TOLD YOU THAT 608 00:31:19,760 --> 00:31:21,880 WE ALSO ARE INTERESTED IN 609 00:31:21,880 --> 00:31:25,440 DEVELOPING BIOLOGICS, THAT CAN 610 00:31:25,440 --> 00:31:27,400 BE USED TO TREAT MULTIPLE 611 00:31:27,400 --> 00:31:27,960 SCLEROSIS. 612 00:31:27,960 --> 00:31:31,880 FOR THESE STUDIES WE USED 613 00:31:31,880 --> 00:31:32,760 EXPERIMENTAL AUTOIMMUNE 614 00:31:32,760 --> 00:31:33,760 ENCEPHALOMYELITIS MODEL, EAE. 615 00:31:33,760 --> 00:31:38,120 WHICH IS A MODEL OF MULTIPLE 616 00:31:38,120 --> 00:31:38,560 SCLEROSIS. 617 00:31:38,560 --> 00:31:44,720 WE INDUCED DISEASE BY IMMUNIZING 618 00:31:44,720 --> 00:31:48,440 MICE WITH MOG PEPTIDES, IN 619 00:31:48,440 --> 00:31:54,120 ADJUVANTS, FOLLOWED MICE FOR 620 00:31:54,120 --> 00:31:55,880 ABOUT 30 DAYS. 621 00:31:55,880 --> 00:32:01,200 EXAMINED ALL THE SYMPTOMS THAT 622 00:32:01,200 --> 00:32:02,640 CHARACTERIZED EAE IN MICE. 623 00:32:02,640 --> 00:32:06,440 AND AS YOU CAN SEE, THESE MICE 624 00:32:06,440 --> 00:32:11,280 HERE WERE PROTECTED FROM SEVERE 625 00:32:11,280 --> 00:32:11,920 EAE. 626 00:32:11,920 --> 00:32:15,760 THE SAME DATA SHOWN HERE BY 627 00:32:15,760 --> 00:32:17,400 HISTOLOGY, THESE BLACK ARROWS 628 00:32:17,400 --> 00:32:18,400 INDICATE AREAS OF INFLAMMATION, 629 00:32:18,400 --> 00:32:21,120 AND AS YOU CAN SEE MICE THAT 630 00:32:21,120 --> 00:32:26,800 WERE THE CONTROL MICE HERE 631 00:32:26,800 --> 00:32:28,240 DEVELOPED -- HAD SIGNIFICANT 632 00:32:28,240 --> 00:32:31,200 INFILTRATION OF CELLS INTO THE 633 00:32:31,200 --> 00:32:37,120 BRAIN AND SPINAL CORD. 634 00:32:37,120 --> 00:32:40,280 AND WE ALSO USED BLUE DYE TO 635 00:32:40,280 --> 00:32:41,920 STAIN THE SECTIONS, AND THIS 636 00:32:41,920 --> 00:32:45,200 ALLOWS US TO DETECT AREAS OF 637 00:32:45,200 --> 00:32:47,160 DEMYELINATION, WHICH IS AN 638 00:32:47,160 --> 00:32:49,800 IMPORTANT ASPECT OF THE DISEASE 639 00:32:49,800 --> 00:32:50,880 IN MULTIPLE SCLEROSIS. 640 00:32:50,880 --> 00:32:56,560 AS YOU CAN SEE, MICE THAT 641 00:32:56,560 --> 00:33:00,520 RECEIVED IL-27 WERE PROTECTED 642 00:33:00,520 --> 00:33:02,160 FROM THESE HALLMARK FEATURES OF 643 00:33:02,160 --> 00:33:03,560 EAE IN MICE. 644 00:33:03,560 --> 00:33:07,400 IN THIS STUDY WHAT WE DID THEN 645 00:33:07,400 --> 00:33:10,560 WAS TO USE WHAT WE CALL TWO 646 00:33:10,560 --> 00:33:14,840 CONGENIC MOUSE STRAINS, ONE IS 647 00:33:14,840 --> 00:33:18,560 CD45.2, ONE IS CD45.1. 648 00:33:18,560 --> 00:33:21,640 WE ISOLATED THE BREGULARS FROM 649 00:33:21,640 --> 00:33:26,240 THE CD45.2 MICE, INDUCED EAE, IN 650 00:33:26,240 --> 00:33:28,640 THESE MICE. 651 00:33:28,640 --> 00:33:32,920 AND THEN TREATED THEM WITH THESE 652 00:33:32,920 --> 00:33:35,640 BREGS, THIS SYSTEM ALLOWS US TO 653 00:33:35,640 --> 00:33:38,920 MONITOR AND TRACE AND FIND OUT 654 00:33:38,920 --> 00:33:42,960 THE SOURCE OF THE BREGS THAT 655 00:33:42,960 --> 00:33:44,320 CONTRIBUTED IN THE SUPPRESSION 656 00:33:44,320 --> 00:33:46,040 OF THE DISEASE. 657 00:33:46,040 --> 00:33:52,040 AND WHAT THIS DATA IS SHOWING 658 00:33:52,040 --> 00:33:56,760 HERE IS THAT BOTH THE DONOR 659 00:33:56,760 --> 00:33:59,920 CELLS THAT WERE THE SOURCE OF 660 00:33:59,920 --> 00:34:01,560 THE BREGS, THEY THEY ALSO EXPAND 661 00:34:01,560 --> 00:34:05,400 IN VIVO, IN THE ANIMALS, 662 00:34:05,400 --> 00:34:07,520 RECIPIENTS, IN ADDITION THE 663 00:34:07,520 --> 00:34:11,080 ANIMALS THAT THE RECIPIENT MICE 664 00:34:11,080 --> 00:34:14,560 ALSO -- NORMAL CELLS WERE ALSO 665 00:34:14,560 --> 00:34:18,200 CONVERTED, TO REGULATORY CELLS 666 00:34:18,200 --> 00:34:23,560 THAT MAKE IL-27 AND IL-10. 667 00:34:23,560 --> 00:34:28,040 AND THE MECHANISM BY WHICH THEY 668 00:34:28,040 --> 00:34:31,560 CONTRIBUTED TO SUPPRESS EAE HERE 669 00:34:31,560 --> 00:34:32,880 IN THIS MODEL. 670 00:34:32,880 --> 00:34:37,480 I WILL NOW SWITCH GEARS AND TELL 671 00:34:37,480 --> 00:34:41,080 YOU ABOUT AN EXOSOME 672 00:34:41,080 --> 00:34:41,840 IMMUNOTHERAPY. 673 00:34:41,840 --> 00:34:44,760 DURING CHARACTERIZATION OF THE 674 00:34:44,760 --> 00:34:48,360 IL-27 BREGS AND IL-35 BREGS WE 675 00:34:48,360 --> 00:34:52,640 DISCOVERED THAT THEY SECRETE 676 00:34:52,640 --> 00:34:57,000 EXOSOMES THAT CONTAIN IL-27 AND 677 00:34:57,000 --> 00:35:05,040 IL-35, WE CALL THESE EXOSOMES 678 00:35:05,040 --> 00:35:08,080 IL-27 AND IL-35 EXOSOMES, THEY 679 00:35:08,080 --> 00:35:10,360 RANGE FROM 50 TO 150 NANOMETERS, 680 00:35:10,360 --> 00:35:12,480 AND THIS ENABLES THEM TO BE ABLE 681 00:35:12,480 --> 00:35:16,520 TO CROSS THE BLOOD-BRAIN BARRIER 682 00:35:16,520 --> 00:35:18,560 AND BLOOD-RETINAL BARRIER MAKING 683 00:35:18,560 --> 00:35:20,440 THEM IDEAL VEHICLES FOR 684 00:35:20,440 --> 00:35:22,320 THERAPEUTIC DELIVERY OF IMMUNE 685 00:35:22,320 --> 00:35:26,000 SUPPRESSIVE CYTOKINES LIKE IL-35 686 00:35:26,000 --> 00:35:29,520 AND IL-27 INTO THE BRAIN OR THE 687 00:35:29,520 --> 00:35:31,400 RETINA. 688 00:35:31,400 --> 00:35:32,920 WE BELIEVE THAT EXOSOMES ARE 689 00:35:32,920 --> 00:35:35,640 MAJOR THERAPIES FOR INFLAMMATORY 690 00:35:35,640 --> 00:35:35,920 DISEASES. 691 00:35:35,920 --> 00:35:37,840 AND I WILL PROVIDE DATA, OUR 692 00:35:37,840 --> 00:35:40,120 CONCLUSION DERIVES FROM THE FACT 693 00:35:40,120 --> 00:35:44,880 THAT WE'RE ABLE TO USE IL-35 694 00:35:44,880 --> 00:35:49,120 EXOSOMES AND IL-27 TO SUPPRESS 695 00:35:49,120 --> 00:35:50,320 UVEITIS AND GRAFT-VERSUS-HOST 696 00:35:50,320 --> 00:35:50,760 DISEASE. 697 00:35:50,760 --> 00:35:55,800 SO IN THIS STUDY WE ISOLATED I 698 00:35:55,800 --> 00:35:59,400 35 EXOSOMES FROM CULTURES FROM 699 00:35:59,400 --> 00:36:09,960 THE SUPER NATES OF CULTURES OF I 700 00:36:13,080 --> 00:36:14,080 35 BREGS, EXTRACELLULAR 701 00:36:14,080 --> 00:36:18,040 VESICLES, USING TO ISOLATE THE 702 00:36:18,040 --> 00:36:22,280 EXOSOMES, AND EXOSOMES RANGE AS 703 00:36:22,280 --> 00:36:26,640 SHOWN FROM 50 TO 150 NANOMETERS. 704 00:36:26,640 --> 00:36:28,760 WE INDUCED EAU IN THESE MICE. 705 00:36:28,760 --> 00:36:33,120 AFTER NINE DAYS STARTED TREATING 706 00:36:33,120 --> 00:36:37,040 THEM WITH -- WE GAVE THEM 20 707 00:36:37,040 --> 00:36:40,000 MILLION OF THE I 35 EXOSOMES 708 00:36:40,000 --> 00:36:40,760 RETRO ORBITALLY, INJECTED MORE 709 00:36:40,760 --> 00:36:42,840 OR LESS IN THE EYE. 710 00:36:42,840 --> 00:36:46,320 AND THEN EVALUATED THE EFFECT OF 711 00:36:46,320 --> 00:36:52,360 THE EXOSOMES BY FUNDUSCOPY, 712 00:36:52,360 --> 00:36:55,120 HISTOLOGY, OCT AND ERG. 713 00:36:55,120 --> 00:36:57,840 AGAIN, IT GAVE RESOUNDING 714 00:36:57,840 --> 00:37:00,440 ANSWER, THAT THE EXOSOMES CAN 715 00:37:00,440 --> 00:37:03,840 SUPPRESS THE UVEITIS IN THESE 716 00:37:03,840 --> 00:37:04,560 MICE. 717 00:37:04,560 --> 00:37:07,480 AND SUPPRESSION DERIVES FROM 718 00:37:07,480 --> 00:37:09,560 INHIBITING PRO-INFLAMMATORY 719 00:37:09,560 --> 00:37:11,400 CYTOKINES AND UPREGULATING 720 00:37:11,400 --> 00:37:16,040 IMMUNOSUPPRESSIVE CYTOKINES AS 721 00:37:16,040 --> 00:37:17,440 DEMONSTRATED BY ELISA. 722 00:37:17,440 --> 00:37:20,800 WE REPEATED THE SAME EXPERIMENT 723 00:37:20,800 --> 00:37:27,920 FOR IL-27 EXOSOMES, WHICH WE SAW 724 00:37:27,920 --> 00:37:31,960 ISOLATED FROM CULTURES OF I 27 725 00:37:31,960 --> 00:37:37,240 BREGS, RECAPITULATED SAME 726 00:37:37,240 --> 00:37:39,080 RESULTS WITH IL-35 EXOSOMES. 727 00:37:39,080 --> 00:37:41,160 THE ADDITIONAL DATA THAT WE GOT 728 00:37:41,160 --> 00:37:44,360 FROM THIS DATA, FROM THIS STUDY, 729 00:37:44,360 --> 00:37:49,600 IS THAT ONE OF THE WAYS THAT THE 730 00:37:49,600 --> 00:37:51,440 EXOSOMES INHIBIT INFLAMMATIONS 731 00:37:51,440 --> 00:37:55,840 IS INDUCING CHECKPOINT 732 00:37:55,840 --> 00:38:00,320 INHIBITORS LIKE PD-1, CTLA-4, 733 00:38:00,320 --> 00:38:03,240 INDUCES INCREASED EXPRESSION IN 734 00:38:03,240 --> 00:38:05,000 T CELLS AND THIS CHECKPOINT 735 00:38:05,000 --> 00:38:06,680 INHIBITORS ARE KNOWN TO INDUCE T 736 00:38:06,680 --> 00:38:09,160 CELL EXHAUSTION AND SO ONE OF 737 00:38:09,160 --> 00:38:12,880 THE WAYS THAT THEY USE 738 00:38:12,880 --> 00:38:14,080 SUPPRESSING THE PATHOGENIC 739 00:38:14,080 --> 00:38:19,120 LYMPHOCYTES IS BY INDUCING T 740 00:38:19,120 --> 00:38:21,000 CELL EXHAUSTION. 741 00:38:21,000 --> 00:38:22,280 I WOULD NOW TURN TO 742 00:38:22,280 --> 00:38:26,480 GRAFT-VERSUS-HOST DISEASE WHICH 743 00:38:26,480 --> 00:38:31,480 IS ANOTHER INFLAMMATORY DISEASE. 744 00:38:31,480 --> 00:38:32,600 745 00:38:32,600 --> 00:38:33,680 GRAFT-VERSUS-HOST DISEASE IS 746 00:38:33,680 --> 00:38:34,880 CHRONIC AUTOIMMUNE DISORDER THAT 747 00:38:34,880 --> 00:38:38,480 CAN AFFECT MOST TISSUES OF THE 748 00:38:38,480 --> 00:38:38,720 BODY. 749 00:38:38,720 --> 00:38:41,360 WE THERE EXAMINED WHETHER WE 750 00:38:41,360 --> 00:38:47,160 COULD USE I 35 BREGS OR EXOSOMES 751 00:38:47,160 --> 00:38:48,000 TO TREAT GRAFT-VERSUS-HOST 752 00:38:48,000 --> 00:38:48,800 DISEASE. 753 00:38:48,800 --> 00:38:53,960 THIS IS A MODEL OF THE GvHD 754 00:38:53,960 --> 00:38:57,040 MODEL WE USED IN THIS STUDY, 755 00:38:57,040 --> 00:39:01,920 CONSISTS OF USING TWO MICE THAT 756 00:39:01,920 --> 00:39:08,200 DIFFER IN THEIR H2K LOCUS. 757 00:39:08,200 --> 00:39:12,080 THE RECIPIENT MICE WE GAVE IT 758 00:39:12,080 --> 00:39:17,440 10.4 GRAY UNIT OF RADIATION, FOR 759 00:39:17,440 --> 00:39:18,360 15 MINUTES. 760 00:39:18,360 --> 00:39:24,200 AND SIX TO EIGHT HOURS BEFORE 761 00:39:24,200 --> 00:39:24,600 TRANSPLANTATION. 762 00:39:24,600 --> 00:39:26,080 FOR THE TRANSPLANTATION, WE USED 763 00:39:26,080 --> 00:39:30,880 DONOR MICE HERE, AND WE 764 00:39:30,880 --> 00:39:31,840 OBTAINED -- WE TRANSPLANTED 10 765 00:39:31,840 --> 00:39:34,480 MILLION T CELL DEPLETED BONE 766 00:39:34,480 --> 00:39:35,360 MARROW CELLS. 767 00:39:35,360 --> 00:39:37,480 FOR THE ANIMAL THAT WOULD HAVE 768 00:39:37,480 --> 00:39:38,320 GRAFT-VERSUS-HOST DISEASE WE 769 00:39:38,320 --> 00:39:42,680 GAVE THEM THE BONE MARROW CELL 770 00:39:42,680 --> 00:39:45,440 AS WELL AS 50 MILLION ALLOGENEIC 771 00:39:45,440 --> 00:39:47,080 SPLEEN CELLS. 772 00:39:47,080 --> 00:39:50,560 FOR THE TREATMENT GROUP, WE GAVE 773 00:39:50,560 --> 00:39:52,440 THEM THE SAME REGIMENT AS 774 00:39:52,440 --> 00:39:55,680 GRAFT-VERSUS-HOST DISEASE BUT 775 00:39:55,680 --> 00:40:03,720 ALSO INCLUDED 1.5 MILLION I35 776 00:40:03,720 --> 00:40:04,200 BREGS. 777 00:40:04,200 --> 00:40:05,360 HISTOLOGICAL EXAMINATION WERE 778 00:40:05,360 --> 00:40:07,240 RESTRICTED TO LUNGS, SPLEEN, 779 00:40:07,240 --> 00:40:10,160 LIVER, COLON. 780 00:40:10,160 --> 00:40:12,960 THIS IS SCORING METHOD TO SCORE 781 00:40:12,960 --> 00:40:13,800 GRAFT-VERSUS-HOST DISEASE, 782 00:40:13,800 --> 00:40:15,440 LOOKED AT BODY WEIGHT CHANGE, 783 00:40:15,440 --> 00:40:22,360 POSTURE, ACTIVITY OF THE MICE, 784 00:40:22,360 --> 00:40:24,200 SKIN INTEGRITY. 785 00:40:24,200 --> 00:40:26,200 AND THE GRAFT-VERSUS-HOST 786 00:40:26,200 --> 00:40:28,040 DISEASE SCORE IS SUBMISSION OF 787 00:40:28,040 --> 00:40:28,960 THESE FIVE PARAMETERS. 788 00:40:28,960 --> 00:40:33,640 AND I WOULD LIKE TO MENTION HERE 789 00:40:33,640 --> 00:40:35,760 THIS WAS A VERY LABOR INTENSIVE 790 00:40:35,760 --> 00:40:38,240 STUDY BECAUSE A LOT OF POSTDOCS 791 00:40:38,240 --> 00:40:40,680 AND STUDENTS HAD TO GO TO THE 792 00:40:40,680 --> 00:40:42,840 ANIMAL ROOM EVERY DAY TO EXAMINE 793 00:40:42,840 --> 00:40:43,680 THESE MICE. 794 00:40:43,680 --> 00:40:47,080 AND IT WAS PARTICULARLY 795 00:40:47,080 --> 00:40:49,960 CHALLENGING DURING THE COVID 796 00:40:49,960 --> 00:40:52,680 PANDEMIC. 797 00:40:52,680 --> 00:40:56,520 NOW, IN THIS MODEL THAT WE USE, 798 00:40:56,520 --> 00:40:58,800 MICE THAT DID NOT RECEIVE ANY 799 00:40:58,800 --> 00:41:00,200 TREATMENT THAT HAD 800 00:41:00,200 --> 00:41:01,000 GRAFT-VERSUS-HOST DISEASE, THEY 801 00:41:01,000 --> 00:41:04,160 DIED WITHIN 45 DAYS. 802 00:41:04,160 --> 00:41:06,040 ALL OF THEM. 803 00:41:06,040 --> 00:41:06,480 100%. 804 00:41:06,480 --> 00:41:11,280 AS SHOWN HERE MICE THAT RECEIVED 805 00:41:11,280 --> 00:41:14,680 IL-35 TREATMENT, THEY SURVIVED 806 00:41:14,680 --> 00:41:16,080 TILL 90 DAYS. 807 00:41:16,080 --> 00:41:19,280 AND HOWEVER IT'S ONLY 60% OF 808 00:41:19,280 --> 00:41:20,600 THEM THAT SURVIVED TO 90 DAYS 809 00:41:20,600 --> 00:41:23,880 BUT IT'S A VERY, VERY 810 00:41:23,880 --> 00:41:26,360 SIGNIFICANT PROGRESS IN THIS 811 00:41:26,360 --> 00:41:28,040 PARTICULAR THERAPEUTIC APPROACH. 812 00:41:28,040 --> 00:41:33,080 AND ALSO, YOU SEE HERE THAT IT 813 00:41:33,080 --> 00:41:35,120 ALSO SUPPRESSED MICE THAT HAS 814 00:41:35,120 --> 00:41:37,880 SUPPRESSION OF CLINICAL SYMPTOMS 815 00:41:37,880 --> 00:41:44,000 OF GRAFT-VERSUS-HOST DISEASE IN 816 00:41:44,000 --> 00:41:50,480 MICE, I 35 BREGS, STUDY TISSUES 817 00:41:50,480 --> 00:41:52,440 FROM THE LIVER, THE COLON, THE 818 00:41:52,440 --> 00:41:56,160 SPLEEN, AND THE BRAIN. 819 00:41:56,160 --> 00:42:01,120 AND WE SENT THIS OUT 820 00:42:01,120 --> 00:42:01,960 COMMERCIALLY FOR UNBIASED 821 00:42:01,960 --> 00:42:03,360 PATHOLOGIST TO SCORE THE 822 00:42:03,360 --> 00:42:03,600 DISEASE. 823 00:42:03,600 --> 00:42:06,120 AND THIS IS THE RESULT THAT THEY 824 00:42:06,120 --> 00:42:08,960 CAME UP WITH. 825 00:42:08,960 --> 00:42:10,600 AND AGAIN, ACROSS THE BOARD, 826 00:42:10,600 --> 00:42:12,800 WHETHER LIVER OR COLON OR SPLEEN 827 00:42:12,800 --> 00:42:16,520 OR LUNG, THE DISEASE IS 828 00:42:16,520 --> 00:42:17,960 SIGNIFICANTLY REDUCED. 829 00:42:17,960 --> 00:42:18,840 AND DISEASE REDUCTION DERIVED 830 00:42:18,840 --> 00:42:24,080 FROM THE FACT THAT THE IL-35 831 00:42:24,080 --> 00:42:25,920 BREGS SUPPRESS CYTOKINES THAT 832 00:42:25,920 --> 00:42:29,440 CAUSE WHAT WE CALL CYTOKINE 833 00:42:29,440 --> 00:42:31,240 STORM, WHICH IS RESPONSIBLE TO A 834 00:42:31,240 --> 00:42:32,800 GREAT EXTENT TO THE PATHOLOGY 835 00:42:32,800 --> 00:42:35,600 THAT'S SEEN IN GRAFT-VERSUS-HOST 836 00:42:35,600 --> 00:42:36,600 DISEASE. 837 00:42:36,600 --> 00:42:39,880 WE LOOKED AT THE CELLS IN 838 00:42:39,880 --> 00:42:40,760 DIFFERENT TISSUES, AND, AGAIN, 839 00:42:40,760 --> 00:42:45,880 IN THE LIVER, WE SEE SIGNIFICANT 840 00:42:45,880 --> 00:42:48,760 EXPANSION OF IL-10-PRODUCING B 841 00:42:48,760 --> 00:42:53,120 CELLS AND IL-35-PRODUCING B 842 00:42:53,120 --> 00:42:57,280 CELLS IN THIS MODEL. 843 00:42:57,280 --> 00:43:03,160 WE REPEATED THIS EXPERIMENT 844 00:43:03,160 --> 00:43:06,240 USING THE I35 EXOSOMES, AGAIN 845 00:43:06,240 --> 00:43:10,520 SHOWING WE -- ALL OF OUR 846 00:43:10,520 --> 00:43:15,440 EXOSOMES, SOME WE VERIFIED USING 847 00:43:15,440 --> 00:43:16,400 ELECTROMICROSCOPY TO SHOW HERE 848 00:43:16,400 --> 00:43:19,280 THEY ALL RANGE IN SIZE BETWEEN 849 00:43:19,280 --> 00:43:23,440 50 AND 150 NANOMETERS. 850 00:43:23,440 --> 00:43:25,840 AGAIN, I 35 EXOSOME PROLONGED 851 00:43:25,840 --> 00:43:27,280 SURVIVAL OF GRAFT-VERSUS-HOST 852 00:43:27,280 --> 00:43:32,280 DISEASE MICE SHOWN IN THIS, 80% 853 00:43:32,280 --> 00:43:35,920 OF THEM SURVIVED, BY DAY 40 MOST 854 00:43:35,920 --> 00:43:39,320 OF THE 45, MOST ALL OF THE 855 00:43:39,320 --> 00:43:40,520 GRAFT-VERSUS-HOST DISEASE THAT 856 00:43:40,520 --> 00:43:43,560 WERE NOT MICE WHO WERE NOT 857 00:43:43,560 --> 00:43:47,000 TREATED DIED. 858 00:43:47,000 --> 00:43:48,200 AGAIN, THE I35 EXOSOMES 859 00:43:48,200 --> 00:43:49,040 EXPRESSED GRAFT-VERSUS-HOST 860 00:43:49,040 --> 00:43:51,880 DISEASE IN THIS MODEL. 861 00:43:51,880 --> 00:43:54,280 AND, AGAIN, DISEASE SUPPRESSION 862 00:43:54,280 --> 00:43:57,800 DERIVED FROM SUPPRESSION OF 863 00:43:57,800 --> 00:44:01,080 CYTOKINES THAT MEDIATE CYTOKINE 864 00:44:01,080 --> 00:44:03,480 STORM, WHICH CONTRIBUTES HEAVILY 865 00:44:03,480 --> 00:44:04,960 FOR THE DISEASE PATHOLOGY. 866 00:44:04,960 --> 00:44:08,200 AGAIN, WHEN WE LOOKED AT THE 867 00:44:08,200 --> 00:44:13,440 CELLS, WE SEE THAT THE T CELLS 868 00:44:13,440 --> 00:44:21,800 IN THESE ANIMALS, THEY EXPRESS 869 00:44:21,800 --> 00:44:26,920 HIGH LEVELS OF CTLA-4, PD-1 AND 870 00:44:26,920 --> 00:44:29,120 CHECKPOINT INHIBITORS PLAY A 871 00:44:29,120 --> 00:44:32,720 ROLE ON THE INTEGRITY OF CELLS, 872 00:44:32,720 --> 00:44:37,680 T CELL EXHAUSTION, REDUCED 873 00:44:37,680 --> 00:44:42,160 EFFECTIVE NUMBER IN VIVO. 874 00:44:42,160 --> 00:44:43,720 I WILL NOW SUMMARIZE. 875 00:44:43,720 --> 00:44:44,880 WHAT THE TAKEHOME MESSAGE FROM 876 00:44:44,880 --> 00:44:47,600 ALL THE STUDIES I TOLD YOU WILL 877 00:44:47,600 --> 00:44:49,280 REGULATORY B CELLS AND THEIR 878 00:44:49,280 --> 00:44:50,760 EXOSOMES, WELL, THE TAKEHOME 879 00:44:50,760 --> 00:44:58,120 MESSAGE IS THAT THE BREGS AND 880 00:44:58,120 --> 00:45:00,320 EXOSOMES THEY PRODUCE, POTENTIAL 881 00:45:00,320 --> 00:45:01,440 EMERGING IMMUNOTHERAPIES FOR 882 00:45:01,440 --> 00:45:06,440 UVEITIS, MULTIPLE SCLEROSIS, AND 883 00:45:06,440 --> 00:45:07,440 GRAFT-VERSUS-HOST DISEASE. 884 00:45:07,440 --> 00:45:15,720 I WOULD END BY SHARING WITH YOU 885 00:45:15,720 --> 00:45:19,600 GAPS IN KNOWLEDGE BETWEEN THESE 886 00:45:19,600 --> 00:45:30,160 AND CYTOKINES IL-35 AND IL-27. 887 00:45:30,920 --> 00:45:34,920 THEY ARE COMPRISED OF TWO 888 00:45:34,920 --> 00:45:35,240 SUBUNITS. 889 00:45:35,240 --> 00:45:36,640 WE KNOW THE GENES THAT CODE, WE 890 00:45:36,640 --> 00:45:38,840 KNOW CELLS THAT PRODUCE THEM, 891 00:45:38,840 --> 00:45:44,360 MOSTLY PRELIMINARY -- LYMPHOCYD 892 00:45:44,360 --> 00:45:48,520 MYELOID CELLS, WE KNOW SOME 893 00:45:48,520 --> 00:45:49,840 REGULATORY PROPERTIES. 894 00:45:49,840 --> 00:46:00,320 EACH OF THE SUBUNIT PROTEINS 895 00:46:00,840 --> 00:46:08,520 ENCODE SIGNAL PEPTIDES, TWO 896 00:46:08,520 --> 00:46:10,520 SUBUNITS THAT MAKE IL-35 AND 897 00:46:10,520 --> 00:46:13,040 IL-27 EVENTUALLY ASSOCIATE VERVE 898 00:46:13,040 --> 00:46:18,160 TO FORM FUNCTIONAL ON IL-27 899 00:46:18,160 --> 00:46:19,480 CYTOKINE. 900 00:46:19,480 --> 00:46:21,480 HOWEVER, NOBODY KNOWS THE 901 00:46:21,480 --> 00:46:24,440 MECHANISM BY WHICH THESE ARE 902 00:46:24,440 --> 00:46:24,680 CAUSED. 903 00:46:24,680 --> 00:46:29,240 AND THE DATA THAT WE HAVE IS 904 00:46:29,240 --> 00:46:31,560 THAT SO CONTRARY TO PREVAILING 905 00:46:31,560 --> 00:46:37,440 VIEW THAT IL-35 AND IL-27 ARE 906 00:46:37,440 --> 00:46:39,400 SECRETED CYTOKINES, I WILL SHOW 907 00:46:39,400 --> 00:46:42,840 YOU DATA THAT THE P 35 IS 908 00:46:42,840 --> 00:46:45,240 TRANSMEMBRANE PROTEIN AND NOT A 909 00:46:45,240 --> 00:46:47,000 SOLUBLE PROTEIN, AND PROVIDE 910 00:46:47,000 --> 00:46:50,360 EVIDENCE THAT THE EBI3 IS THE 911 00:46:50,360 --> 00:46:54,680 ONLY ONE CYTOPLASMIC OR SECRETED 912 00:46:54,680 --> 00:46:56,160 PROTEIN AND SHOW EARLIER RECENT 913 00:46:56,160 --> 00:47:03,520 DATA THAT IN RESPONSE TO INTENSE 914 00:47:03,520 --> 00:47:05,280 INFLAMMATION, IL-35 -- EBI 3, 915 00:47:05,280 --> 00:47:07,600 GOES TO THE MEMBRANE, CYCLES 916 00:47:07,600 --> 00:47:11,920 BACK, ASSOCIATES WITH P 35 TO 917 00:47:11,920 --> 00:47:13,920 FORM MEMBRANE BOUND IL-35 918 00:47:13,920 --> 00:47:14,280 CYTOKINES. 919 00:47:14,280 --> 00:47:17,080 SO IL-35 WE DON'T BELIEVE IS 920 00:47:17,080 --> 00:47:18,640 SOLUBLE PROTEIN. 921 00:47:18,640 --> 00:47:21,680 WE BELIEVE IT'S MEMBRANE BOUND 922 00:47:21,680 --> 00:47:22,280 CYTOKINE. 923 00:47:22,280 --> 00:47:26,320 THIS DATA HERE, WE USE 924 00:47:26,320 --> 00:47:29,080 COMBINATION OF 925 00:47:29,080 --> 00:47:29,720 IMMUNOHISTOCHEMISTRY AND 926 00:47:29,720 --> 00:47:31,240 CONFOCAL MICROSCOPY, AND THESE 927 00:47:31,240 --> 00:47:32,200 ANALYSIS WE ACTIVATED THESE 928 00:47:32,200 --> 00:47:35,440 CELLS AND AS YOU CAN SEE HERE IN 929 00:47:35,440 --> 00:47:37,720 THE PLASMA MEMBRANE HERE IN 930 00:47:37,720 --> 00:47:40,400 GREEN, YOU SEE THE P 35. 931 00:47:40,400 --> 00:47:44,360 IT'S NOT IN THE CYTOPLASM. 932 00:47:44,360 --> 00:47:48,760 AND THIS BLUE AREA HERE IS JUST 933 00:47:48,760 --> 00:47:51,440 DEPICTS THE NUCLEUS, THE DYE. 934 00:47:51,440 --> 00:47:52,840 WHEN YOU LOOK AT EPI 3 AS YOU 935 00:47:52,840 --> 00:47:55,880 SEE IT'S NOT ON THE MEMBRANE, 936 00:47:55,880 --> 00:48:01,040 IT'S MOSTLY IN THE CYTOPLASM. 937 00:48:01,040 --> 00:48:04,600 AND I'M SHOWING YOU THIS PROTEIN 938 00:48:04,600 --> 00:48:06,120 HERE. 939 00:48:06,120 --> 00:48:15,320 IT'S A CD8 CD81 PROTEIN, SIGNAG 940 00:48:15,320 --> 00:48:20,600 PROTEINS TO PLASMA MEMBRANE 941 00:48:20,600 --> 00:48:22,640 PARTICULARLY CD19 PROTEIN. 942 00:48:22,640 --> 00:48:24,040 INTO THE PLASMA MEMBRANE. 943 00:48:24,040 --> 00:48:29,680 WHAT WE SHOW HERE IS THAT THE P 944 00:48:29,680 --> 00:48:34,120 35 CO-LOCALIZES WITH CD81, MAYBE 945 00:48:34,120 --> 00:48:36,560 CD81 PLAYS A ROLE IN TRAFFICKING 946 00:48:36,560 --> 00:48:39,520 THE P 35 TO THE PLASMA MEMBRANE 947 00:48:39,520 --> 00:48:49,200 AND AGAIN WHEN YOU LOOK AT THE 948 00:48:49,200 --> 00:48:54,920 IMAGE HERE EBI 3 STAYS IN THE 949 00:48:54,920 --> 00:48:56,480 CYTOPLASM, RETICULUM, SHOWING 950 00:48:56,480 --> 00:49:00,400 THE INTERACTION BETWEEN 81 AND P 951 00:49:00,400 --> 00:49:03,360 35 HERE BUT PROXIMITY LIGATION 952 00:49:03,360 --> 00:49:07,200 ASSAY THAT INTERACT. 953 00:49:07,200 --> 00:49:09,480 HOWEVER, DURING PROLONGED 954 00:49:09,480 --> 00:49:11,120 ACTIVATION OF THESE CELLS, AND 955 00:49:11,120 --> 00:49:13,960 THIS DATA WAS GENERATED BY A 956 00:49:13,960 --> 00:49:15,920 GRADUATE STUDENT WHO JUST 957 00:49:15,920 --> 00:49:20,640 RECEIVED A Ph.D. THIS PAST 958 00:49:20,640 --> 00:49:24,040 YEAR, AND WHAT WE SEE IS THAT 959 00:49:24,040 --> 00:49:26,640 THE EBI3 THEN EITHER MIGRATES 960 00:49:26,640 --> 00:49:29,200 FROM THE CYTOPLASM OR 961 00:49:29,200 --> 00:49:30,480 TRANSPORTED FROM THE CYTOPLASM 962 00:49:30,480 --> 00:49:33,680 TO THE PLASMA MEMBRANE OR FROM 963 00:49:33,680 --> 00:49:36,920 THE CYTOSOL, AND FORMS 964 00:49:36,920 --> 00:49:42,200 TRIMOLECULAR COMPLEX WITH CD81, 965 00:49:42,200 --> 00:49:45,000 P35, AND EBI3 TO FORM MEMBRANE 966 00:49:45,000 --> 00:49:46,560 BOUND IL-35. 967 00:49:46,560 --> 00:49:50,520 WE DID THE SIMILAR THING WITH 968 00:49:50,520 --> 00:49:54,440 THE IL-27 BREGS, AS YOU CAN 969 00:49:54,440 --> 00:49:58,600 APPRECIATE HERE THE EBI - 28 IS 970 00:49:58,600 --> 00:50:01,760 FOUND IN THE PLASMA MEMBRANE, TO 971 00:50:01,760 --> 00:50:09,680 SOME EXTENT IN CYTO CYTOPLASM. 972 00:50:09,680 --> 00:50:11,640 EBI 3 IS MOSTLY CYTOPLASMIC. 973 00:50:11,640 --> 00:50:16,800 WHEN YOU DO THE IMAGE HERE YOU 974 00:50:16,800 --> 00:50:20,960 SEE CO-LOCALIZATION OF CD81 AND 975 00:50:20,960 --> 00:50:21,840 P28. 976 00:50:21,840 --> 00:50:26,960 MEANWHILE EBI3 IS MOSTLY SO 977 00:50:26,960 --> 00:50:29,480 CYTOPLASMIC, SHOWN BETTER HERE. 978 00:50:29,480 --> 00:50:39,120 IN THIS STUDY HERE WE DID ONE 979 00:50:39,120 --> 00:50:49,640 ADDITIONAL THING, LABELED ALSO 980 00:50:52,960 --> 00:50:59,960 DETECT CALRETICULIN IN CYTOPLASM 981 00:50:59,960 --> 00:51:02,560 AND ENDOPLASMIC RETICULUM. 982 00:51:02,560 --> 00:51:04,880 I WILL STOP HERE AND INTRODUCE 983 00:51:04,880 --> 00:51:06,680 YOU TO SOME OF THE VERY HAPPY 984 00:51:06,680 --> 00:51:08,320 PEOPLE IN MY LAB WHO DID ALL 985 00:51:08,320 --> 00:51:14,040 THIS WORK THAT I DESCRIBED HERE 986 00:51:14,040 --> 00:51:14,800 THIS AFTERNOON. 987 00:51:14,800 --> 00:51:16,200 I WOULD LIKE TO ACKNOWLEDGE MY 988 00:51:16,200 --> 00:51:26,720 COLLEAGUES AT THE EYE INSTITUTE, 989 00:51:27,120 --> 00:51:28,600 PARTICULARLY THE ONE WHO 990 00:51:28,600 --> 00:51:31,640 RECRUITED ME AS A NEWLY MINTED 991 00:51:31,640 --> 00:51:33,920 Ph.D. TO WORK WITH HIM. 992 00:51:33,920 --> 00:51:37,640 I'VE BEEN HERE IN THE NEI ON THE 993 00:51:37,640 --> 00:51:39,720 NIH SINCE THEN. 994 00:51:39,720 --> 00:51:42,040 THIS IS THE ONLY JOB I'VE HAD AS 995 00:51:42,040 --> 00:51:43,040 AN ADULT. 996 00:51:43,040 --> 00:51:48,680 AND I LOVE WORKING AT NIH. 997 00:51:48,680 --> 00:51:54,720 AND MY COLLEAGUES, RACHEL AND 998 00:51:54,720 --> 00:51:56,000 CHI-CHAO-CHAN, COLLABORATORS AND 999 00:51:56,000 --> 00:51:59,080 FRIENDS, WE PUBLISHED SEVERAL 1000 00:51:59,080 --> 00:51:59,840 PAPERS TOGETHER, AND HAVE 1001 00:51:59,840 --> 00:52:02,800 ENJOYED WORKING WITH THEM ALL 1002 00:52:02,800 --> 00:52:10,680 THESE YEARS AT NIH. 1003 00:52:10,680 --> 00:52:13,080 ROBERT NUSSENBLATT WAS AT BRANCH 1004 00:52:13,080 --> 00:52:14,960 LAB CHIEF WHO DIED A FEW YEARS 1005 00:52:14,960 --> 00:52:22,360 AGO, WE STILL MISS HIM. 1006 00:52:22,360 --> 00:52:25,240 YINGYOU AND MARY HAVE BEEN -- 1007 00:52:25,240 --> 00:52:27,280 WE'VE COLLABORATED ON MANY 1008 00:52:27,280 --> 00:52:28,400 MANUSCRIPTS, THEY PLAY IMPORTANT 1009 00:52:28,400 --> 00:52:31,560 ROLE IN SOME OF OUR ANIMAL MODEL 1010 00:52:31,560 --> 00:52:32,520 MAL STUDIES. 1011 00:52:32,520 --> 00:52:34,520 RAFAEL IS THE HEAD OF THE FLOW 1012 00:52:34,520 --> 00:52:40,200 CYTOMETRY SCORE AND HE'S BEEN AN 1013 00:52:40,200 --> 00:52:42,840 INVALUABLE MEMBER AND HELPED US 1014 00:52:42,840 --> 00:52:51,440 IN MANY OF OUR LYMPHOCYTE 1015 00:52:51,440 --> 00:52:53,880 STUDIES. 1016 00:52:53,880 --> 00:52:54,760 LIJIN AND GUANPUA PLAYED 1017 00:52:54,760 --> 00:52:56,960 IMPORTANT ROLE TO GENERATE MICE 1018 00:52:56,960 --> 00:53:00,120 WITH TARGETED DELETION OF IL-27 1019 00:53:00,120 --> 00:53:03,080 RECEPTOR FROM B CELLS. 1020 00:53:03,080 --> 00:53:07,360 AND MONIKA AND YURI, THEY PLAYED 1021 00:53:07,360 --> 00:53:10,960 SEMINAL IMPORTANT ROLE IN HPLC 1022 00:53:10,960 --> 00:53:19,600 PURIFICATION OF THE IL-35 1023 00:53:19,600 --> 00:53:20,680 CYTOKINE. 1024 00:53:20,680 --> 00:53:22,080 ROBERT AND MONES, THEY HELPED US 1025 00:53:22,080 --> 00:53:26,840 A LOT WITH THE CONFOCAL 1026 00:53:26,840 --> 00:53:30,960 MICROSCOPY AND THE E.M. IMAGE 1027 00:53:30,960 --> 00:53:33,280 WAS GENERATED BY MONIS. 1028 00:53:33,280 --> 00:53:38,080 PAUL WINGFIELD HELPED US WITH 1029 00:53:38,080 --> 00:53:39,840 CHARACTERIZING THE HETERODIMERIC 1030 00:53:39,840 --> 00:53:42,360 IL-35 TO DEMONSTRATE IT'S 1031 00:53:42,360 --> 00:53:44,960 HETERODIMERIC PROTEIN. 1032 00:53:44,960 --> 00:53:48,920 LAST BUT NOT LEAST, MICHELE K 1033 00:53:48,920 --> 00:53:54,200 EVANS, SENIOR INVESTIGATOR AT 1034 00:53:54,200 --> 00:53:54,880 NATIONAL AGING INSTITUTE, WHO 1035 00:53:54,880 --> 00:53:56,160 HAPPENED TO BE MY WIFE. 1036 00:53:56,160 --> 00:54:00,880 SHE'S THE ONE WHO INTRODUCED ME 1037 00:54:00,880 --> 00:54:05,880 TO THE WONDERFUL FIELD OF 1038 00:54:05,880 --> 00:54:07,000 EXOSOMES. 1039 00:54:07,000 --> 00:54:13,320 AND THIS OCCURRED ONE EVENING 1040 00:54:13,320 --> 00:54:16,840 DURING DINNER, COMPLAINING ABOUT 1041 00:54:16,840 --> 00:54:20,240 THE IL-27, IL-35 BREGS WITH 1042 00:54:20,240 --> 00:54:21,000 SUBUNITS, THERAPEUTICALLY 1043 00:54:21,000 --> 00:54:23,000 DIFFICULT TO BE ABLE TO PREDICT 1044 00:54:23,000 --> 00:54:26,920 HOW MUCH OF THEM FORM 1045 00:54:26,920 --> 00:54:27,240 HETERODIMER. 1046 00:54:27,240 --> 00:54:32,240 AND IN VIVO. 1047 00:54:32,240 --> 00:54:34,640 THE THERAPY. 1048 00:54:34,640 --> 00:54:38,080 SHE ASKED HAVE YOU TRIED TO MAKE 1049 00:54:38,080 --> 00:54:38,320 EXOSOMES? 1050 00:54:38,320 --> 00:54:39,480 THEY MAY SIMPLIFY YOUR LIFE. 1051 00:54:39,480 --> 00:54:40,880 JUST DOING THAT EXPERIMENT 1052 00:54:40,880 --> 00:54:43,240 THAT'S HOW COME WE CAME TO FIND 1053 00:54:43,240 --> 00:54:44,960 OUT THEY MAKE EXOSOMES AND IT'S 1054 00:54:44,960 --> 00:54:46,600 OPENED A WONDERFUL AREA OF 1055 00:54:46,600 --> 00:54:48,040 RESEARCH IN MY LAB. 1056 00:54:48,040 --> 00:54:50,360 THANK YOU ALL VERY, VERY MUCH 1057 00:54:50,360 --> 00:54:50,880 FOR LISTENING. 1058 00:54:50,880 --> 00:54:52,320 AND I'LL BE HAPPY TO TAKE ANY 1059 00:54:52,320 --> 00:54:56,360 QUESTIONS IF WE HAVE TIME. 1060 00:54:56,360 --> 00:54:59,600 >> THANK YOU, DR. EGWUAGU, FOR 1061 00:54:59,600 --> 00:55:01,480 SUCH A WONDERFUL TALK ON THE 1062 00:55:01,480 --> 00:55:05,840 STORY OF THE IL-35 AND IL-27 AND 1063 00:55:05,840 --> 00:55:06,360 EXOSOMES. 1064 00:55:06,360 --> 00:55:07,200 THIS IS FANTASTIC. 1065 00:55:07,200 --> 00:55:09,840 WE DO HAVE SOME QUESTIONS. 1066 00:55:09,840 --> 00:55:12,880 LET ME START FIRST. 1067 00:55:12,880 --> 00:55:15,720 I GUESS THE IL-35 STORY THAT YOU 1068 00:55:15,720 --> 00:55:19,520 STARTED OUT WITH, YOU KNOW, VERY 1069 00:55:19,520 --> 00:55:20,080 INTRIGUING. 1070 00:55:20,080 --> 00:55:24,920 DO YOU THINK THAT THERAPIES 1071 00:55:24,920 --> 00:55:28,600 COULD BE USED, USING IL-35 AND 1072 00:55:28,600 --> 00:55:31,360 TOWARDS OTHER DISEASES LIKE 1073 00:55:31,360 --> 00:55:33,080 MACULAR DEGENERATION OR DIABETIC 1074 00:55:33,080 --> 00:55:37,400 RETINOPATHY, OTHER TYPES OF 1075 00:55:37,400 --> 00:55:37,640 DISEASES? 1076 00:55:37,640 --> 00:55:39,120 >> YES, I DO. 1077 00:55:39,120 --> 00:55:45,360 THEY CAN BE APPLIED TO MOST 1078 00:55:45,360 --> 00:55:47,000 INFLAMMATORY DISEASES. 1079 00:55:47,000 --> 00:55:48,960 HOWEVER, MY OUR PREDICTION IS 1080 00:55:48,960 --> 00:55:52,240 THAT WE MAY BE BETTER OFF USING 1081 00:55:52,240 --> 00:55:55,120 EXOSOMES BECAUSE THEY HAVE MORE 1082 00:55:55,120 --> 00:55:55,640 ACCESS. 1083 00:55:55,640 --> 00:55:57,880 THEY CAN EASILY ENTER DIFFERENT 1084 00:55:57,880 --> 00:56:02,560 TISSUES, ESPECIALLY FOR LIKE 1085 00:56:02,560 --> 00:56:03,840 MACULAR DEGENERATION, YOU 1086 00:56:03,840 --> 00:56:07,440 COULD -- THEY HAVE ACCESS TO THE 1087 00:56:07,440 --> 00:56:12,400 CNS AND MUCH MORE USEFUL. 1088 00:56:12,400 --> 00:56:14,880 BASICALLY IS THE ACTIVE CYTOKINE 1089 00:56:14,880 --> 00:56:25,480 ITSELF, SO THE BREGS OR EXOSOMS 1090 00:56:30,760 --> 00:56:31,760 ON VECTORS. 1091 00:56:31,760 --> 00:56:35,720 I BELIEVE THEY CAN BE APPLICABLE 1092 00:56:35,720 --> 00:56:38,320 TO MANY INFLAMMATORY DISEASE. 1093 00:56:38,320 --> 00:56:42,440 HOWEVER, THEY SHOULD BE 1094 00:56:42,440 --> 00:56:44,520 INHIBITED IN CANCER, BECAUSE 1095 00:56:44,520 --> 00:56:47,040 THEY WOULD SUPPRESS CANCER 1096 00:56:47,040 --> 00:56:47,400 IMMUNOTHERAPY. 1097 00:56:47,400 --> 00:56:49,600 >> VERY INTERESTING. 1098 00:56:49,600 --> 00:56:50,240 THANK YOU. 1099 00:56:50,240 --> 00:56:51,880 WE HAVE A QUESTION THAT WAS 1100 00:56:51,880 --> 00:56:56,240 E-MAILED FROM ONE OF OUR NHLBI 1101 00:56:56,240 --> 00:56:58,120 COLLEAGUES. 1102 00:56:58,120 --> 00:57:00,760 COULD I 27, I 35 B CELL 1103 00:57:00,760 --> 00:57:03,480 TREATMENTS BE HELPFUL IN 1104 00:57:03,480 --> 00:57:04,760 TREATING DANGEROUS CYTOKINE 1105 00:57:04,760 --> 00:57:06,520 STORMS WITH CAR T CELL TREATMENT 1106 00:57:06,520 --> 00:57:06,960 IN THE FUTURE? 1107 00:57:06,960 --> 00:57:08,400 >> I DON'T SEE WHY NOT. 1108 00:57:08,400 --> 00:57:11,880 BASICALLY WHAT IT DOES IS THAT 1109 00:57:11,880 --> 00:57:14,840 IT'S ABLE TO INHIBIT LYMPHOCYTES 1110 00:57:14,840 --> 00:57:18,440 THAT MAKE THOSE CYTOKINES THAT 1111 00:57:18,440 --> 00:57:20,160 INDUCE, AND IT'S ACROSS THE 1112 00:57:20,160 --> 00:57:20,480 BOARD. 1113 00:57:20,480 --> 00:57:23,840 SO THAT IT'S NOT JUST LIKE ONE 1114 00:57:23,840 --> 00:57:26,040 CYTOKINE INHIBITS A REPERTOIRE 1115 00:57:26,040 --> 00:57:31,920 OF CYTOKINES, WHETHER IT'S IL-1, 1116 00:57:31,920 --> 00:57:33,560 INTERFERON GAMMA, LOOKED AT TYPE 1117 00:57:33,560 --> 00:57:35,320 I INTERFERONS BUT INTERFERON 1118 00:57:35,320 --> 00:57:37,960 GAMMA AND TNF AND ALL THE OTHER 1119 00:57:37,960 --> 00:57:41,360 CYTOKINES, YES, THEY CAN INHIBIT 1120 00:57:41,360 --> 00:57:41,560 THEM. 1121 00:57:41,560 --> 00:57:41,720 YES. 1122 00:57:41,720 --> 00:57:45,000 >> GREAT. 1123 00:57:45,000 --> 00:57:45,240 THANK YOU. 1124 00:57:45,240 --> 00:57:46,920 I HAVE ANOTHER QUESTION. 1125 00:57:46,920 --> 00:57:50,840 YOU DESCRIBED A LOT TO US TODAY 1126 00:57:50,840 --> 00:57:52,840 RELATED TO THE I 35 SORT OF 1127 00:57:52,840 --> 00:57:54,280 STORY, IF YOU WILL. 1128 00:57:54,280 --> 00:57:57,000 IS THERE MORE FOR THE IL-27 1129 00:57:57,000 --> 00:57:58,880 STORY THAT MAY BE FORTHCOMING? 1130 00:57:58,880 --> 00:58:02,560 OR DID ONE PATH JUST LEAD MORE 1131 00:58:02,560 --> 00:58:02,960 TOWARDS THE IL-35? 1132 00:58:02,960 --> 00:58:05,760 >> AS A MATTER OF FACT, THE 1133 00:58:05,760 --> 00:58:12,640 IL-27 IS MORE INTRIGUING BECAUSE 1134 00:58:12,640 --> 00:58:14,480 IT'S AN INNATE-LIKE LYMPHOCYTE, 1135 00:58:14,480 --> 00:58:23,160 AND WHILE IL-27 CAN INDUCE IL-35 1136 00:58:23,160 --> 00:58:25,800 AND IL-10, IL-35 CANNOT INDUCE 1137 00:58:25,800 --> 00:58:27,760 IL-27. 1138 00:58:27,760 --> 00:58:27,960 OKAY. 1139 00:58:27,960 --> 00:58:29,080 SO IT HAS A MUCH MORE ROBUST 1140 00:58:29,080 --> 00:58:29,680 ACTIVITY. 1141 00:58:29,680 --> 00:58:33,880 THE OTHER THING I DIDN'T POINT 1142 00:58:33,880 --> 00:58:37,440 OUT, A LOT OF BREGS, THEY ARE 1143 00:58:37,440 --> 00:58:39,720 SPECIFIC TO PARTICULAR DISEASES. 1144 00:58:39,720 --> 00:58:43,960 IL-27 CAN BE ELICITED JUST USING 1145 00:58:43,960 --> 00:58:44,720 LPS. 1146 00:58:44,720 --> 00:58:48,200 SO THE IL-27 WE GENERATE TO 1147 00:58:48,200 --> 00:58:50,920 INHIBIT EAE WE COULD USE IT TO 1148 00:58:50,920 --> 00:58:51,800 INHIBIT EAU. 1149 00:58:51,800 --> 00:58:54,760 SO, IT HAS A MUCH MORE GLOBAL 1150 00:58:54,760 --> 00:58:55,000 IMPACT. 1151 00:58:55,000 --> 00:58:59,080 AND PART OF IT IS BECAUSE IT'S 1152 00:58:59,080 --> 00:59:02,280 REALLY AN INNATE-LIKE CELL, 1153 00:59:02,280 --> 00:59:09,400 MEANWHILE THE IL-35 BREGS IS 1154 00:59:09,400 --> 00:59:10,840 ADAPTED BRREG REALIZE MORE RICH, 1155 00:59:10,840 --> 00:59:16,960 YOU COULD USE IT TO INHIBIT 1156 00:59:16,960 --> 00:59:20,160 DIVERSE AUTOIMMUNE DISEASES. 1157 00:59:20,160 --> 00:59:23,840 >>THANK YOU FOR SUCH A 1158 00:59:23,840 --> 00:59:24,840 WONDERFUL TALK AND TAKING TIME 1159 00:59:24,840 --> 00:59:25,960 TO ANSWER QUESTIONS FROM THE 1160 00:59:25,960 --> 00:59:26,400 AUDIENCE. 1161 00:59:26,400 --> 00:59:27,600 WE'RE AT 1:00. 1162 00:59:27,600 --> 00:59:29,120 THIS WILL CONCLUDE OUR SESSION 1163 00:59:29,120 --> 00:59:29,640 FOR GRAND ROUNDS. 1164 00:59:29,640 --> 00:59:30,440 THANK YOU VERY MUCH. 1165 00:59:30,440 --> 00:59:31,400 AND THANK YOU TO THE AUDIENCE 1166 00:59:31,400 --> 00:59:32,000 FOR YOUR LISTENING AND YOUR 1167 00:59:32,000 --> 00:00:00,000 PARTICIPATION.