Natural history of MELAS associated with mitochondrial DNA m.3243A>G genotype.

OBJECTIVE: To describe the natural history of clinical and laboratory features associated with the m.3243A>G mitochondrial DNA point mutation. Natural history data are needed to obtain prognostic information and for clinical trial planning. METHODS: We included 85 matrilineal relatives from 35 families with at least 2 visits in this prospective cohort study. Thirty-one were fully symptomatic with mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS), and 54 were carrier relatives. Evaluations included standardized questionnaires (medical history and daily living functioning), physical examination, neuropsychological testing, and a battery of imaging and laboratory tests. We evaluated changes in clinical and laboratory features over time and survival. Outcomes are reported over a follow-up period of up to 10.6 years (mean 3.8 ± 2.2 years for patients and 5.5 ± 3.0 for carrier relatives). RESULTS: Neurologic examination, neuropsychological testing, and daily living scores significantly declined in all patients with MELAS, whereas no significant deterioration occurred in carrier relatives. Cerebral MRI scores declined significantly in patients with MELAS. Magnetic resonance spectroscopy estimates of lactate in the lateral ventricles increased over time, and high lactate was associated with increased mortality. Symptom onset in childhood often was associated with worse outcome. Patients with MELAS had a greater death rate than carrier relatives. CONCLUSIONS: Patients with MELAS carrying the m.3243A>G mutation show a measurable decline in clinical and imaging outcomes. It is hoped that these data will be helpful in anticipating the disease course and in planning clinical trials for MELAS.

Dichloroacetate causes toxic neuropathy in MELAS: a randomized, controlled clinical trial.

OBJECTIVE: To evaluate the efficacy of dichloroacetate (DCA) in the treatment of mitochondrial myopathy, encephalopathy, lactic acidosis and stroke-like episodes (MELAS). BACKGROUND: High levels of ventricular lactate, the brain spectroscopic signature of MELAS, correlate with more severe neurologic impairment. The authors hypothesized that chronic cerebral lactic acidosis exacerbates neuronal injury in MELAS and therefore, investigated DCA, a potent lactate-lowering agent, as potential treatment for MELAS. METHODS: The authors conducted a double-blind, placebo-controlled, randomized, 3-year cross-over trial of DCA (25 mg/kg/day) in 30 patients (aged 10 to 60 years) with MELAS and the A3243G mutation. Primary outcome measure was a Global Assessment of Treatment Efficacy (GATE) score based on a health-related event inventory, and on neurologic, neuropsychological, and daily living functioning. Biologic outcome measures included venous, CSF, and 1H MRSI-estimated brain lactate. Blood tests and nerve conduction studies were performed to monitor safety. RESULTS: During the initial 24-month treatment period, 15 of 15 patients randomized to DCA were taken off study medication, compared to 4 of 15 patients randomized to placebo. Study medication was discontinued in 17 of 19 patients because of onset or worsening of peripheral neuropathy. The clinical trial was terminated early because of peripheral nerve toxicity. The mean GATE score was not significantly different between treatment arms. CONCLUSION: DCA at 25 mg/kg/day is associated with peripheral nerve toxicity resulting in a high rate of medication discontinuation and early study termination. Under these experimental conditions, the authors were unable to detect any beneficial effect. The findings show that DCA-associated neuropathy overshadows the assessment of any potential benefit in MELAS.

[Type 1 glucose transporter (Glut1) deficiency: manifestations of a hereditary neurological syndrome]. / Deficiencia del transportador de glucosa tipo 1 (Glut1): manifestaciones de un síndrome neurológico hereditario.

AIM: To define this genetic syndrome. DEVELOPMENT: The constellation of infantile epilepsy, acquired microcephaly and hypoglychorrachia is characteristic of glucose transporter type 1 (Glut1) deficiency syndrome, a prototype neurometabolic disorder caused by inheritable mutations in the gene SLC2A1. All known mutations reduce the function of Glut1 in the blood brain barrier and thus limit brain glucose availability. As the cerebral metabolic rate for glucose increases during infancy, patients become gradually symptomatic, a phenomenon that underscores the importance of early diagnosis via lumbar puncture and treatment, which has meet with some success in ameliorating several --but not all-- features of the disease. CONCLUSION: The increasing number of mild phenotypic variants being described, owing to the improved awareness of the disease, has led to the consideration of Glut1 deficiency in the diagnosis of infantile seizures, mental retardation, familial epilepsy and movement disorders.

Deficiencia del transportador de glucosa tipo 1 (Glut1): manifestaciones de un síndrome neurológico hereditario / Type 1 glucose transporter (Glut1) deficiency: manifestations of a hereditary neurological syndrome

Objetivo. Definir este síndrome genético. Desarrollo. El síndrome de deficiencia del transportador de glucosa tipo 1 (Glut1), prototipo de enfermedad neurometabólica, combina manifestaciones como la epilepsia infantil, la microcefalia adquirida y la hipoglucorraquia, debidas a mutación del gen SLC2A1. Todas las mutaciones conocidas en SLC2A1 reducen la función del trasportador Glut1 en la barrera hematoencefálica, y limitan la captación de glucosa por el sistema nervioso. A medida que la tasa de consumo cerebral de glucosa aumenta durante la infancia, los pacientes acusan más la sintomatología, lo que justifica la necesidad de establecer el diagnóstico en las etapas iniciales de la enfermedad mediante la punción lumbar y el comienzo del tratamiento lo antes posible. Este último consigue mejorar varias manifestaciones graves del síndrome, aunque no todas ellas, por ahora. Conclusiones.El conocimiento de la enfermedad, junto con el de la existencia de variantes fenotípicas leves o incompletas, han conducido a la consideración de la deficiencia de Glut1 en el estudio de las convulsiones infantiles, el retraso mental, la epilepsia familiar y los trastornos del movimiento (AU)

Aim. To define this genetic syndrome. Development. The constellation of infantile epilepsy, acquired microcephaly and hypoglychorrachia is characteristic of glucose transporter type 1 (Glut1) deficiency syndrome, a prototype neurometabolic disorder caused by inheritable mutations in the gene SLC2A1. All known mutations reduce the function of Glut1 in the blood brain barrier and thus limit brain glucose availability. As the cerebral metabolic rate for glucose increases during infancy, patients become gradually symptomatic, a phenomenon that underscores the importance of early diagnosis via lumbar puncture and treatment, which has meet with some success in ameliorating several –but not all– features of the disease. Conclusion. The increasing number of mild phenotypic variants being described, owing to the improved awareness of the disease, has led to the consideration of Glut1 deficiency in the diagnosis of infantile seizures, mental retardation, familial epilepsy and movement disorders (AU)

Cerebral lactic acidosis correlates with neurological impairment in MELAS.

OBJECTIVE: To evaluate the role of chronic cerebral lactic acidosis in mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS). METHODS: The authors studied 91 individuals from 34 families with MELAS and the A3243G point mutation and 15 individuals from two families with myoclonus epilepsy and ragged red fibers (MERRF) and the A8344G mutation. Subjects were divided into four groups. Paternal relatives were studied as controls (Group 1). The maternally related subjects were divided clinically into three groups: asymptomatic (no clinical evidence of neurologic disease) (Group 2), oligosymptomatic (neurologic symptoms but without the full clinical picture of MELAS or MERRF) (Group 3), and symptomatic (fulfilling MELAS or MERRF criteria) (Group 4). The authors performed a standardized neurologic examination, neuropsychological testing, MRS, and leukocyte DNA analysis in all subjects. RESULTS: The symptomatic and oligosymptomatic MELAS subjects had significantly higher ventricular lactate than the other groups. There was a significant correlation between degree of neuropsychological and neurologic impairment and cerebral lactic acidosis as estimated by ventricular MRS lactate levels. CONCLUSIONS: High levels of ventricular lactate, the brain spectroscopic signature of MELAS, are associated with more severe neurologic impairment.

Role of reentry of in vivo alloMHC peptide-activated T cells into the adult thymus in acquired systemic tolerance.

BACKGROUND: T cell recognition of alloMHC peptide presented by self dendritic cells via the indirect pathway of allorecognition in the thymus induces T cell tolerance. Most recently we have shown that the i.v. administration of immunodominant Wistar Furth MHC class I (RT1.Au) peptide 5- (P5) pulsed myeloid or lymphoid dendritic cells induces operational tolerance to a fully MHC-mismatched cardiac allograft. This finding led us to hypothesize that circulation of peripheral P5-activated T cells to the thymus plays an important role in the induction of acquired tolerance. METHODS: We used the adoptive transfer of 111Indium-oxine- (111In-oxine) labeled P5-pulsed syngeneic dendritic cells and in vivo P5-activated syngeneic T cells to study the role of their circulation to the thymus in the induction of transplantation tolerance. RESULTS: Intravenously administered 111In-oxine-labeled naïve DC actively migrated to and localized in the liver and spleen but did not enter the lymph nodes, bone marrow, and thymus. In vitro peptide-pulsed dendritic cells had a similar pattern of tissue localization except for a modest number of myeloid but not lymphoid DC entering the thymus. The demonstration that adoptive transfer of in vivo peptide-primed T cells induces permanent graft survival in antilymphocyte serum transiently immunosuppressed syngeneic secondary hosts led us to examine the traffic of in vivo activated T cells. Whereas naïve syngeneic T cells preferentially homed to the peripheral lymphoid organs, they did not reenter the thymus. In contrast, in vivo peptide-activated peripheral T cells migrated to and accumulated in the thymus, thus confirming that reentry of T cells to the thymus is restricted to in vivo activated T cells. Although antilymphocyte serum immunosuppression significantly reduced circulation of primed T cells to the thymus, it did not completely abolish it, as seen with gamma-irradiated primed T cells. CONCLUSION: These findings provide the first formal evidence directly linking reentry of in vivo alloMHC peptide-activated T cells to the thymus with the induction and possibly maintenance of acquired antigen-specific tolerance. Our results suggest that the thymus is open to a two-way traffic with the periphery and may function as a repository of immunological memory.

Mechanisms of acquired thymic tolerance: induction of transplant tolerance by adoptive transfer of in vivo allomhc peptide activated syngeneic T cells.

BACKGROUND: Our most recent observation that i.v. injection of Wistar-Furth (WF) major histocompatibility complex Class I peptide 5 (P5)-pulsed self-myeloid or lymphoid dendritic cells (DC) induces transplantation tolerance suggests that adoptive transfer of in vivo allopeptide-primed host T cells might induce acquired tolerance through their interaction with thymic DC. METHODS: To examine this hypothesis, host myeloid DC cultured in rat granulocyte/macrophage colony stimulating factor and interleukin 4 were pulsed in vitro with P5 and injected intravenously into syngeneic ACI rats. The T cells primed to P5 via the indirect pathway of allorecognition were harvested 7 days later and administered by either intravenously or intrathymically into syngeneic ACI recipients of WF cardiac allografts. RESULTS: Syngeneic T cells obtained from the spleen of P5-primed rats had a high mixed lymphocyte reaction proliferative response to P5 presented by self-DC. I.v. administration of 2x107 P5-primed alloreactive purified host splenic T cells alone on day -7 significantly (P<0.001) prolonged cardiac allograft survival from 10.5+/-1.0 days to 18.6+/-1.8 days in the WF-to-ACI rat combination. I.v. injection of P5-activated host T cells combined with 0.5 ml antilymphocyte serum (ALS)-transient immunosuppression on day -7 led to 100% donor-specific permanent graft survival (>200 days). Thymectomy before i.v. injection of P5-activated syngeneic T cells led to acute graft rejection, suggesting that the homing of in vivo activated T cells to the host thymus might play a role in the induction of tolerance. To further define the role of the recipient thymus in this model, we examined the effects of intrathymic (i.t.) injection of P5-primed alloreactive T cells on graft survival and found that i.t. administration of the P5-primed T cells on day -7 alone significantly prolonged graft survival (15.0+/-0.7 days) and when combined with 0.5 ml ALS led to donor-specific permanent graft survival. The long-term unresponsive recipients permanently (>100 days) accepted second-set donor-specific cardiac allografts but not third-party (Lewis) grafts. CONCLUSIONS: These findings demonstrate that the adoptive transfer of splenic T cells primed to an indirectly presented donor peptide induces transplantation tolerance in a transiently immunosuppressed secondary syngeneic recipient. Our data suggest that the interaction of thymic DC with activated peripheral T cells induces alloantigen (Ag)-specific T-cell tolerance by either inactivation or deletion of alloreactive T cells in the thymus. This observation provides the first formal evidence that the interaction between thymic DC and activated peripheral T cells that continuously circulate through the thymus plays an important role in the induction and maintenance of Ag-specific tolerance.

[Fatalities caused by anticholinergic antiparkinsonian drugs. Analysis of findings in a 11-year national material]. / Dødsfall forårsaket av antikolinerge antiparkinsonmedikamenter. Analysefunn i et nasjonalt 11-årsmateriale.

All autopsy samples received at the National Institute of Forensic Toxicology during the years 1986-1996 which contained anticholinergic antiparkinsonian drugs were reviewed. Of a total of 69 cases, orphenadrine was present in 57 (83%), biperiden in 8 (12%), procyclidine in 3 (4%), and trihexyphenidyl/benzhexol in 1 (1%) of the subjects. The measured concentrations were assessed in light of previously published data. Of 21 cases where causality between drug ingestion and death was classified as either highly probable (18/21) or possible (3/21), all subjects tested positive for orphenadrine. In the autopsy samples from these patients, orphenadrine concentrations in the 4.5-600 mumol/l range (mean 62.5 mumol/l, SD 126.5 mumol/l) were determined. Because of a low national autopsy rate, there is reason to believe that the actual numbers of drug-related deaths in this period may have been significantly higher. It is concluded that orphenadrine is responsible for a disproportionally high number of overdose deaths.

Induction of peripheral tolerance by intrathymic inoculation of soluble alloantigens: evidence for the role of host antigen-presenting cells and suppressor cell mechanism.

Intrathymic (IT) inoculation of soluble alloantigens (Ag) obtained from 3 M KCl extracts of resting T-cells induces donor-specific tolerance to cardiac allografts and islet allografts. This study examined the cellular basis of induction of transplantation tolerance by IT injection of soluble Ag. Our results show that while IT inoculation of 2 mg soluble donor Ag on Day -7 relative to Lewis islet transplantation induced specific unresponsiveness to islet allografts (> 200 days) in naive diabetic recipients, IT inoculation of 2 mg soluble Ag on the same day as islet transplantation did not prolong islet allograft survival in the same Lewis-to-WF rat combination. To define the role of donor APCs in intrathymic tolerance, we showed that IT injection of an admixture of 1 x 10(4) donor DC and 2 mg soluble Ag caused acute islet graft rejection. In contrast, addition of 1 x 10(4) recipient-type DC to the IT inoculum did not prevent long-term graft survival. This finding suggests that while the presence of donor APCs in the inoculum does not appear necessary for IT-alloantigen to induce peripheral tolerance, presentation of the soluble Ag in the thymus is dependent on host APCs. This conclusion is supported by our in vitro MLR experiments which showed that in vivo WF-Ag-primed Lewis T-cells proliferated specifically to WF-soluble Ag and that the response was enhanced 14-fold by the addition of responder-type DC. Addition of anti-Lewis MHC class II mAb specifically blocked the alloresponse, thus suggesting that in vivo Ag-primed T-cells are capable of recognizing and proliferating in response to allopeptides presented by responder APCs. We also showed that adoptive transfer of syngeneic naive T-cells into unresponsive recipients failed to break tolerance to long-term surviving islet allografts. This finding suggests that tolerance in this model is not due to a lack of T help. On the other hand, the adoptive transfer of spleen cells, but not sera, from the unresponsive WF recipients bearing long-term (> 120 days) functioning Lewis islets resulted in prolonged survival of donor-type but not third-party islet allografts in secondary syngeneic hosts. Our data suggest that the tolerogenic effect of IT inoculation of soluble Ag is dependent on the indirect pathway of Ag presentation and clonal deletion of alloreactive T-cells in the thymus, while suppressor/regulatory mechanism may be involved in the maintenance of peripheral tolerance.

Prevention of graft-versus-host disease and bone marrow rejection: kinetics of induction of tolerance by UVB modulation of accessory cells and T cells in the bone marrow inoculum.

UVB irradiation (700 J/m2) of bone marrow cells (UVB-BMC) before transplantation into lethally gamma-irradiated (10.5 Gy) allogeneic rats prevents graft-versus-host disease (GVHD) and induces a stable complete lymphohematopoietic chimerism. To better understand the underlying mechanism of the development of stable chimerism and induction of tolerance to donor organs in this model, we examined if the addition of T cells or dendritic cells (DC), as antigen presenting cells (APC), would restore the immunogenicity of UVB-BMC in in vitro mixed lymphocyte reaction (MLR) and induce in vivo bone marrow (BM) graft rejection. Whereas gamma-irradiated, unfractionated BMC induce allogeneic T cells to proliferate, UVB irradiation of BMC abolishes the stimulatory capacity of such cells in a primary MLR. Addition of purified T cells, CD4+ T cells, CD8+ T cells or B cells, respectively, failed to restore the capacity of UVB-BMC to stimulate allogeneic T-cell proliferation. In contrast, the addition of only a small number of splenic accessory cells or purified DC, which by themselves were relatively ineffective in stimulating T-cell proliferation, restored the accessory function and the allostimulatory capacity of UVB-BMC. To define the molecular defect induced by UVB irradiation, cytokines were added as costimulatory factors to primary MLRs and the results showed that the addition of interleukin (IL)-2 or IL-6 but not IL-1 or interferon gamma (IFN-gamma) restored the stimulatory capacity of UVB BMC. This finding suggests that UVB may alter the production, and/or utilization of IL-2 and IL-6 either at the membrane or cytoplasmic level. Parallel in vivo studies showed that addition of DC to UVB BM inoculum resulted in failure of BM engraftment, whereas addition of T cells led to development of fatal GVHD, thus suggesting that UVB modulation of accessory cells reduces graft immunogenicity and prevents BMT rejection, while modulation of T cells prevents GVHD. Our data provide evidence that UVB modulation of APC and mature T cells contained within BMC is potentially useful in preventing GVHD without endangering successful engraftment and may serve as a model for induction of adult chimerism and tolerance without the development of GVHD.

Induction of stable chimerism and transplantation tolerance to rat islet and heart allografts by ultraviolet-B modulation of bone marrow cells.

Ultraviolet-B irradiation (UV-B) (700 J/m2) of BM cells prior to transplantation into lethally gamma-irradiated (1050 rads) allogeneic rats prevents the development of GVHD and results in stable chimerism. This study was developed to determine if UV-B modulation of BMT is useful for preconditioning recipients for the induction of tolerance to donor islets and heart allografts. Lethally irradiated Lewis rats that received UV-B irradiated (700 J/m2) WF BMT (10(8) BM cells) demonstrated stable chimerism without any evidence of GVHD. The stable Lewis chimeras were made diabetic with streptozotocin (STZ) at 28-35 days after BMT and subdivided into 3 experimental groups that received 1000-1200 islets from WF, Lewis, or BN (third-party), respectively. The results showed that group I diabetic Lewis chimeras accepted permanently (greater than 300 days) BM donor WF islets and became normoglycemic. When 3 of 6 Lewis chimeras transplanted with WF islets were rechallenged with WF hearts 60 days after islet grafts, they accepted both islets and cardiac allografts permanently (greater than 240 days). Similarly, the remaining 3 animals accepted Lewis cardiac allografts permanently, thus indicating tolerance to both donor and recipient alloantigens. Group II diabetic chimeras accepted permanently (greater than 300 days) recipient (Lewis) islets. In contrast, group III chimeras rejected acutely (7-8 days) third-party (BN) islets. However, when these animals that rejected BN islets and again became diabetic were retransplanted with BM donor-type (WF) islets, they became permanently normoglycemic (greater than 200 days). This finding emphasizes the specificity of the induction of tolerance in this model and the apparent lack of organ-specific sensitization. To define the underlying mechanism of tolerance, in vivo adoptive transfer of 10(8) spleen cells to naive Lewis or WF recipients, obtained from tolerant Lewis chimeras carrying donor islets and heart allografts, showed no prolongation of cardiac allografts in the unmodified syngeneic hosts, thus questioning the role of suppressor mechanisms in the tolerant rats. Furthermore, cells from the tolerant chimeras that showed no mixed lymphocyte reaction (MLR) response to Lewis or; WF alloantigens failed to suppress anti-Lewis and anti-WF MLR-response in coculture MLR. These results suggest that tolerance to donor alloantigens in the UV-B BMT model is most likely due to selective elimination of anti-BM donor helper or effector cell precursors (clonal deletion) rather than induction of suppressor cell activity. This study demonstrates that this relatively simple and effective approach to modulation of T cells in BM treatment may be potentially useful in the induction of tolerance to donor organs.

Migration patterns of dendritic cells in the rat: comparison of the effects of gamma and UV-B irradiation on the migration of dendritic cells and Lymphocytes.

To further define the underlying mechanisms of immune suppression induced by UV-B irradiation, we have examined the kinetics of homing patterns of in vitro UV-B-irradiated and gamma-irradiated-thoracic duct lymphocytes (TDL) compared to dendritic cells (DC). Our findings show that 111In-oxine-labeled TDL specifically home to the spleen, liver, lymph nodes, and bone marrow with subsequent recirculation of a large number of cells from the spleen to lymph nodes. In contrast, DC preferentially migrate to the spleen and liver with a relatively insignificant distribution to lymph nodes and an absence of subsequent recirculation. Splenectomy prior to cell injection significantly diverts the spleen-seeking DC to the liver but not to the lymph nodes, while the homing of TDL to lymph nodes is significantly increased. In vitro exposure of 111In-oxine labeled TDL to gamma irradiation does not significantly impair immediate homing to lymphoid tissues but inhibits cell recirculation between 3 and 24 hr. In contrast, gamma irradiation does not affect the tissue distribution of labeled DC, suggesting that DC are more radioresistant to gamma irradiation than TDL. Unlike the findings in animals injected with gamma-irradiated cells, UV-B irradiation virtually abolished the homing of TDL to lymph nodes and significantly reduced the homing of the spleen-seeking DC to the splenic compartment while a large number of cells were sequestered in the liver. The results of in vitro cell binding assay show that TDL, unlike DC, have the capacity to bind to high endothelial venules (HEV) within lymph node frozen sections while gamma and UV-B irradiation significantly inhibit the binding of TDL to lymph node HEV. These findings suggest that: (i) DC, unlike TDL, are unable to recirculate from blood to lymph nodes through HEV; (ii) although gamma irradiation impairs TDL recirculation, it does not affect DC tissue distribution; and (iii) UV-B irradiation impairs both TDL and DC migration patterns. We conclude that the lack of capacity of irradiated TDL to home to lymph nodes is due to damage to cell surface homing receptors and that the failure of DC to home to the lymph node microenvironment is related to the absence of HEV homing receptors on their cell surface.

Prevention of GVHD by modulation of rat bone marrow with UV-B irradiation: II. Kinetics of migration of UV-B-irradiated bone marrow cells in naive and lethally irradiated rats.

UV-B irradiation (700 J/m2) of bone marrow (BM) cells prior to transplantation into lethally gamma-irradiated (1050 rad) allogeneic rats prevents the development of GVHD and results in a stable mixed lymphohematopoietic chimerism. To better understand the underlying mechanisms of the development of stable radiation chimeras in this model, this study was designed to examine whether the dose (700 J/m2) of UV-B irradiation used for the modulation of the BM inoculum would affect the homing pattern of radiolabeled BM cells compared to that of thoracic duct lymphocytes (TDL) in the naive and lethally irradiated recipients. The results showed that intravenously administered, 111Indium-oxine-labeled, unmodified TDL home specifically to the spleen, lymph nodes, and BM compartments with a subsequent recirculation of a large number of cells from the spleen to the lymph nodes. In contrast, radiolabeled, unmodified BM cells migrate specifically to the spleen, liver, and BM with the lymph nodes, thymus, and nonlymphoid organs containing very little amounts of radioactivity. The stable concentrations of radioactivity in the lymphoid and nonlymphoid compartments between 3 and 72 hr after injection suggest that BM cells, unlike TDL, do not recirculate. The migration pattern of BM cells in the naive recipient was not significantly different from that seen in lethally irradiated animals except for the higher concentration of radioactivity in the spleen and BM of irradiated animals compared to that seen in naive recipients. The similarity of tissue localization of BM cells in naive or in irradiated syngeneic recipients to that of allogeneic recipients suggests that the homing of BM cells is not MHC restricted. Our findings of similarity between tissue localization of UV-B-irradiated labeled BM cells and unmodified BM cells in naive and lethally irradiated recipients suggest that a dose of 700 J/m2 of UV-B irradiation is not capable of impairing BM cell migration although it is sufficient to abolish the homing of TDL to the HEV-bearing organs. Thus, our results show that BM cells are less susceptible to cell damage by UV-B irradiation than lymphocytes thereby providing the rationale for ex vivo modulation (rather than elimination) of mature T-lymphocytes in the donor BM inoculum with UV-B irradiation. This relatively simple and effective approach to modulation of T-cells in donor BM inoculum may be potentially useful in preventing GVHD without endangering successful engraftment in larger animals and in man.

Interstitial class II-positive cell depletion by donor pretreatment with gamma irradiation. Evidence of differential immunogenicity between vascularized cardiac allografts and islets.

We used an allograft pretreatment regimen involving lethal total-body gamma irradiation of donor rats eight days prior to transplantation to compare directly the immunogenicty of immediately-vascularized heart allografts with isolated, neovascularized pancreatic islet allografts. TBI pretreatment of heart donors (IHT) caused a modest prolongation in cardiac allograft survival compared with controls in a low-responding fully-allogeneic strain combination, Lewis-to-ACI. The addition of 10(5) donor-type dendritic cells (DCs) at the time of transplantation reversed the prolongation of IHT in this strain combination. Donor pretreatment with TBI did not lead to prolonged cardiac allograft survival in either a high-responding combination, ACI-to-Lewis, or in an MHC class II-compatible strain combination, F344-to-Lewis. In contrast, islets from irradiated donors (IIT) showed markedly prolonged survival in both low-responding (Lewis-to-ACI) and high-responding (W/F-to-Lewis) strain combinations. The addition of 10(5) donor-type DCs to the islets at the time of transplantation caused the rejection of only two of six IIT in the Lewis-to-ACI combination. Combined pretreatment regimens involving TBI-pretreated cardiac allografts--such as recipient immunosuppression with peritransplant cyclosporine (10 mg/kg on days 0, 1, and 2) or the combination of TBI (1000 cGy on day -3) and cyclophosphamide (300 mg/kg on day -5) in a pretreatment regimen--did not lead to synergistic graft prolongation in the low-responding Lewis-to-ACI combination. Immunohistologic studies showed that eight days after TBI rat hearts and islet were both greater than 90% depleted of class II (0X6+) interstitial dendritic cells while class I (0X18+) expression was unchanged. IIT are able to increase class II expression when donors are treated with recombinant gamma interferon (4000 U/day on days -3, -2, and -1). MLR data showed that ACI recipients of Lewis IIT reacted normally to donor (Lewis) and third-party (W/F) stimulator cells greater than 100 days after transplantation. The apparent greater immunogenicity of heart allografts as compared with islets in this model could be due to (1) a quantitative difference between the number of residual class II-positive cells in hearts and islets or a greater nnon-MHC antigenic load in the heart grafts given its larger size, or (2) a quanlitative difference between hearts and islets due to the presence of a vascular endothelium in the immediately vascularized heart that can present alloantigen and provide a major stimulus to rejection.