The Association of Suppressed Hypoxia-Inducible Factor-1 Transactivation of Angiogenesis With Defective Recovery From Cerebral Ischemic Injury in Aged Rats.

Elderly patients suffer more brain damage in comparison with young patients from the same ischemic stroke. The present study was undertaken to test the hypothesis that suppressed hypoxia-inducible factor-1 (HIF-1) transcription activity is responsible for defective recovery after ischemic stroke in the elders. Aged and young rats underwent 1-h transient middle cerebral artery occlusion (MCAO) to produce cerebral ischemic injury. The initial cerebral infarct volume in the young gradually declined as time elapsed, but in the aged rats remained the same. The defective recovery in the aged was associated with depressed angiogenesis and retarded neurorestoration. There was no difference in HIF-1α accumulation in the brain between the two age groups, but the expression of HIF-1 regulated genes involved in cerebral recovery was suppressed in the aged. In confirmation, inhibition of HIF-1 transactivation of gene expression in the young suppressed cerebral recovery from MCAO as the same as that observed in the aged rats. Furthermore, a copper metabolism MURR domain 1 (COMMD1) was significantly elevated after MCAO only in the brain of aged rats, and suppression of COMMD1 by siRNA targeting COMMD1 restored HIF-1 transactivation and improved recovery from MCAO-induced damage in the aged brain. These results demonstrate that impaired HIF-1 transcription activity, due at least partially to overexpression of COMMD1, is associated with the defective cerebral recovery from ischemic stroke in the aged rats.

Effects of Adoptive Transfer of Tolerogenic Dendritic Cells on Allograft Survival in Organ Transplantation Models: An Overview of Systematic Reviews.

Objective. To dissect the efficacy of Tol-DC therapy with or without IS in multiple animal models of transplantation. Methods and Results. PubMed, Medline, Embase, and the Cochrane Library were searched for reviews published up to April 2015. Six systematic reviews and a total of 61 articles were finally included. Data were grouped by organ transplantation models and applied to meta-analysis. Our meta-analysis shows that Tol-DC therapy successfully prolonged allograft survival to varying extents in all except the islet transplantation models and with IS drugs further prolonged the survival of heart, skin, and islet allografts in mice, but not of heart allografts in rats. Compared with IS drugs alone, Tol-DC therapy with IS extended islet allograft survival in rats but failed to influence the survival of skin, small intestine, and heart allografts in rats or of heart and skin allografts in mice. Conclusion. Tol-DC therapy significantly prolonged multiple allograft survival and further prolonged survival with IS. However, standardized protocols for modification of Tol-DC should be established before its application in clinic.

Systematic review with meta-analysis: HIF-1α attenuates liver ischemia-reperfusion injury.

Ischemia-reperfusion injury (IRI) induces inevitable complications in liver transplantation. Many studies have demonstrated that hypoxia-inducible factor 1α (HIF-1α) plays an important role in IRI. However, the mechanism of its pleiotropic effect remains unclear. This systematic review provides a comprehensive evaluation of all available evidence concerning the function of HIF-1α in transplant-induced hepatic IRI. Data were obtained through a search of Medline (PubMed), Embase, and the Cochrane Library literature review on the effect of HIF-1α in IRI (from inception to 12/2014). RevMan was used to calculate standardized mean difference (SMD) and 95% confidence intervals (CIs). Forty articles met inclusion criteria with 2 clinical and 38 basic studies. Two clinical trials (n = 68) revealed ischemic preconditioning (IPC) aroused protection after hepatic IRI based on the higher level of HIF-1α in IPC group compared with control group. In vitro studies confirmed the salutary effect of IPC disappearance in the inhabitation of stabilized HIF-1α. In vivo animal studies showed different HIF-1α expression and distribution patterns in the ischemia and reperfusion stage due to distinctive partial oxygen pressure gradient intra-liver, and 5 animal studies (n=66) showed that stabilized HIF-1α treatment was associated with lower alanine aminotransferase (ALT) (SMD = -1.58; 95% CI =- 2.65, -0.52) when compared with unstabilized HIF-1α group. Not only decreased liver IR injury, stabilized HIF-1α during the acute phase of IR could also promote graft regeneration capacity leading to better initial function and survival rate. More rigorous studies are needed to gauge the effectiveness due to insufficient sample size and possible publication bias.

An improved technique for cerebrospinal fluid collection of cisterna magna in Rhesus monkeys.

BACKGROUND: Currently-used cerebellomedullary cistern puncture method for collecting cerebrospinal fluid (CSF) from monkeys is simple, inexpensive, and practical, but with high risk for brainstem injury and CSF blood contamination. An improved technique was thus developed and characterized. METHOD: Magnetic resonance imaging was used to identify the space and position of the cisterna magna in monkeys. Accordingly, a newly defined procedure for needle punctuation was tested in comparison with the traditional method. Blood contamination in CSF samples and brainstem injury were determined to define the superior of the improved method over the transitional method. RESULTS: The cisterna magna in monkeys was found to be a "▽" shape. The needle was punctured into the cisterna magna aiming at the wider superior gap avoided brainstem injury. The improved method showed that the rate of blood contamination in the CSF samples was reduced from 66.7% to 16.7%, the higher rate of blood contamination was associated with higher risk for brainstem injury. COMPARISON WITH EXISTING METHODS: In traditional method, the needle is punctured aiming at the inferior gap with high density of blood vessels. In improved method, the needle is punctured aiming at the superior gap, pointing to the nose root while advancing the needle and avoiding injury to blood vessels. CONCLUSIONS: This improved technique not only avoids blood contamination of CSF, but also prevents brainstem injury during the process of CSF collection. It is recommended for adaptation for CSF collection in monkeys.

Over-expression of microRNA-494 up-regulates hypoxia-inducible factor-1 alpha expression via PI3K/Akt pathway and protects against hypoxia-induced apoptosis.

BACKGROUND: Hypoxia-inducible factor-1 alpha (HIF-1α) is one of the key regulators of hypoxia/ischemia. MicroRNA-494 (miR-494) had cardioprotective effects against ischemia/reperfusion (I/R)-induced injury, but its functional relationship with HIF-1α was unknown. This study was undertaken to determine if miR-494 was involved in the induction of HIF-1α. RESULTS: Quantitative RT-PCR showed that miR-494 was up-regulated to peak after 4 hours of hypoxia in human liver cell line L02. To investigate the role of miR-494, cells were transfected with miR-494 mimic or miR-negative control, followed by incubation under normoxia or hypoxia. Our results indicated that overexpression of miR-494 significantly induced the expression of p-Akt, HIF-1α and HO-1 determined by qRT-PCR and western blot under normoxia and hypoxia, compared to negative control (p < 0.05). While LY294002 treatment markedly abolished miR-494-inducing Akt activation, HIF-1α and HO-1 increase under both normoxic and hypoxic conditions (p < 0.05). Moreover, apoptosis detection using Annexin V indicated that overexpression of miR-494 significantly decreased hypoxia-induced apoptosis in L02 cells, compared to control (p < 0.05). MiR-494 overexpression also decreased caspase-3/7 activity by 1.27-fold under hypoxia in L02 cells. CONCLUSIONS: Overexpression of miR-494 upregulated HIF-1α expression through activating PI3K/Akt pathway under both normoxia and hypoxia, and had protective effects against hypoxia-induced apoptosis in L02 cells. Thus, these findings suggested that miR-494 might be a target of therapy for hepatic hypoxia/ischemia injury.

Adoptive infusion of tolerance dendritic cells prolongs survival of small intestine allografts in rats: systematic review and meta-analysis.

BACKGROUND: Postoperative infections and rejection are the main limiting factors of small intestine allograft survival. In this study, we performed a systematic review and meta-analysis to review rat small intestine allograft survival following infusion of tolerance dendritic cells (Tol-DCs) induced by different methods. METHODS: Relevant publications were searched from PubMed database and EMbase database. Meta-analysis was performed using RevMan 5.0 software. We chose allograft survival, mixed leukocyte reaction, Th1/Th2 differentiation, Treg induction, and cytotoxic T lymphocyte activity as the outcomes by which to examine possible mechanisms that promote survival. RESULTS: Eleven suitable articles were identified and assessed. Tol-DCs induced by four methods all prolonged allograft survival. The difference in survival time between the Tol-DC group and the control group was indicated by SMD as follows: drug intervention (SMD = 3.02, 95% CI 1.16 to 4.88, P = 0.001), gene modification (SMD = 2.43, 95% CI 1.77 to 3.10, P < 0.00001), imDC (SMD = 1.76, 95% CI 0.90 to 2.62, P < 0.0001), cytokine induction (SMD = 1.68, 95% CI 0.40 to 2.96, P = 0.01). Tol-DCs were also synergistic with immunosuppressive drugs or costimulation inhibitors, but no immune tolerance was observed. A single-dose intravenous injection of 5×10(6) to 6×10(6) Tol-DCs showed the highest allograft survival. Possible mechanisms included donor-specific T-cell hyporesponsiveness and Th2 differentiation. CONCLUSIONS: Our results demonstrated that Tol-DCs induced by four methods prolong rat small intestine allograft survival. Intravenous infusion of 5×10(6) to 6×10(6) Tol-DCs was the optimum dose in rat small intestine transplantation. Immunosuppressive or costimulatory blockade was synergistic with Tol-DC on graft survival. Additional high-quality studies with larger sample sizes are needed to better investigate small intestinal graft longer term survival.

Adoptive transfusion of tolerance dendritic cells prolongs the survival of skin allografts in mice: a systematic review.

OBJECTIVE: We aim to systematically review adoptive transfusion of tolerogenic dendritic cells (Tol-DCs) induced by different ways to affect skin allograft survival in mice. METHODS: We searched PubMed and EMbase for relevant studies and evaluated the quality of included ones. Taking skin allograft survival time as endpoint outcome, we displayed outcomes of each group using one forest map and dissected possible mechanisms underlying survival prolongation. RESULTS: We included 21 studies, which reported four methods of inducing Tol-DCs with different extents of average allograft survival prolongation: skin allograft survival time was prolonged (the drug intervention group, 63.08 ± 42.92 days, 4.6 folds to control; the cytokine induction group: 26.17 ± 16.20 days, 1.8 folds; the gene modification group: 14.65 ± 17.89 days, 1.5 folds; other derivation group 9.63 ± 24.38 days, 0.5 fold). Possible mechanisms underlying survival prolongation included induction of donor-specific T cell hyporesponsiveness, reduction of cytotoxicity against allografts, Th0 skewing to Th2, and generation or expansion of Treg. Infusion of Tol-DCs in combination with immunosuppressive agents or costimulatory blockade contributed to longer prolongation. Compared to MiHA mismatch, MHCI/II mismatch was a much more important factor to cause skin allograft rejection. CONCLUSION: For MHC or MiHA mismatched, allogeneic skin transplants inbred recipients, adoptive transfusion of Tol-DCs induced by 4 methods prolong skin allograft survival to different extents. Drug intervened Tol-DCs works best. Immunosuppressive agents and/or co-stimulatory blockade contribute to better outcomes. Yet more rigorous studies with larger sample size are needed and more attention to mechanisms should be paid.

Adoptive infusion of tolerogenic dendritic cells prolongs the survival of pancreatic islet allografts: a systematic review of 13 mouse and rat studies.

OBJECTIVE: The first Phase I study of autologous tolerogenic dendritic cells (Tol-DCs) in Type 1 diabetes (T1D) patients was recently completed. Pancreatic islet transplantation is an effective therapy for T1D, and infusion of Tol-DCs can control diabetes development while promoting graft survival. In this study, we aim to systematically review islet allograft survival following infusion of Tol-DCs induced by different methods, to better understand the mechanisms that mediate this process. METHODS: We searched PubMed and Embase (from inception to February 29(th), 2012) for relevant publications. Data were extracted and quality was assessed by two independent reviewers. We semiquantitatively analyzed the effects of Tol-DCs on islet allograft survival using mixed leukocyte reaction, Th1/Th2 differentiation, Treg induction, and cytotoxic T lymphocyte activity as mechanisms related-outcomes. We discussed the results with respect to possible mechanisms that promote survival. RESULTS: Thirteen articles were included. The effects of Tol-DCs induced by five methods on allograft survival were different. Survival by each method was prolonged as follows: allopeptide-pulsed Tol-DCs (42.14 ± 44 days), drug intervention (39 days), mesenchymal stem cell induction (23 days), genetic modification (8.99 ± 4.75 days), and other derivation (2.61 ± 6.98 days). The results indicate that Tol-DC dose and injection influenced graft survival. Single-dose injections of 10(4) Tol-DCs were the most effective for allograft survival, and multiple injections were not superior. Tol-DCs were also synergistic with immunosuppressive drugs or costimulation inhibitors. Possible mechanisms include donor specific T cell hyporesponsiveness, Th2 differentiation, Treg induction, cytotoxicity against allograft reduction, and chimerism induction. CONCLUSIONS: Tol-DCs induced by five methods prolong MHC mismatched islet allograft survival to different degrees, but allopeptide-pulsed host DCs perform the best. Immunosuppressive or costimulatory blockade are synergistic with Tol-DC on graft survival. Multiple injections are not superior to single injection. Yet more rigorously designed studies with larger sample sizes are still needed in future.

Functional analysis of alloreactive memory CD4+ T cells derived from skin transplantation recipient and naïve CD4+ T cells derived from untreated mice.

BACKGROUND: Memory T cells (T(M)s) exhibit differential susceptibility to many immunomodulatory strategies that induce immunologic tolerance in naïve T cells, which are believed to be an important barrier to inhibiting rejection and inducing tolerance. As skin grafts are a common model for acquiring T(M)s, we evaluated function of T(M)s derived from skin grafts. We also assessed the modulatory effects on memory T cells function of the microRNAs miR-155 and miR-181a, which are involved in regulating cytokine secretion and TCR sensitivity to antigen, respectively. METHODS: Memory CD4(+) T cells derived from skin-graft recipient mice, and naïve CD4(+) T cells from untreated mice, were isolated by negative magnetic selection, and then stimulated with dendritic cells pulsed with donor-specific antigens. Effector function and regulating mechanisms were assessed. RESULTS: In contrast to naïve CD4(+) T cells, CD4(+) T(M)s stimulated with donor-specific antigen could quickly generate effector function in terms of proliferation and cytokine secretion; miR-155 and miR-181a levels in CD4(+) T(M)s rapidly increased during immune response compared to naïve CD4(+) T cells. CONCLUSION: Memory CD4(+) T cells derived from skin grafts could be used as an experimental tool for evaluating effects of different immune-modulating strategies on T(M)s. Levels of miR-155 and miR-181a up-regulated quickly in T(M)s, which could be an important mechanism by which T(M)s mediate immune responses rapidly.

Dendritic cells transduced with lentiviral vector targeting RelB gene using RNA interference induce hyporesponsiveness in memory CD4(+) T cells and naïve CD4(+) T cells.

The ability of DCs to induce immune tolerance depends on its maturation status. RelB plays a pivotal role in DCs differentiation. A therapeutic protocol of DCs-based not only induces hyporesponsiveness in T(N)s, but also in alloreactive T(M)s is required. Thus, it is urgent to assess modulatory effects of RelB-silenced DCs on T(M)s and T(N)s. In this study, we constructed lentiviral vector which could efficiently silenced the RelB in DCs (DCs-miR RelB) to keep them immature. These DCs induced antigen-specific hyporesponsiveness in CD4(+) T(N)s. In contrast, upon re-stimulation with mature DCs, CD4(+) T(M)s primed by DCs-miR RelB maintained hyporesponsiveness in terms of proliferation and cytokine production. And these may be associated with micro155 and micro181a expression levels in T(M)s and T(N)s. These results may help developing the DCs-based therapeutical protocols by inducing hyporesponsiveness in CD4(+) T(N)s and T(M)s.

Effects of octreotide on activated pancreatic stellate cell-induced pancreas graft fibrosis in rats.

BACKGROUND: Of solid organ transplantations, pancreas transplantation is associated with the highest incidence of pancreatic fibrosis in the early post-transplantation period. Activated pancreatic stellate cells (PSCs) are the main source of pancreatic fibrosis. Octreotide is widely used as a prophylactic for postoperative complications in pancreas transplant recipients. Recent studies have shown that it can inhibit liver fibrosis. This study investigated the effect of octreotide in pancreas graft fibrosis in rats. MATERIALS AND METHODS: Isolated PSCs from Sprague Dawley rats were co-cultured with different doses of octreotide (1.25, 2.5, 5, 10, 20, and 40 ng/mL). PSC proliferation was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide at 48, 72, and 96 h. The α-smooth muscle actin (α-SMA) and collagen I expressions of PSCs were detected by immunohistochemistry and reverse-transcriptase polymerase chain reaction. Rat heterotopic pancreaticoduodenal transplantation was performed with and without octreotide treatment (0.01 mg/kg). Pancreas grafts were harvested at postoperative d 1, 3, 5, and 7. Hematoxylin-eosin staining, Masson's trichrome staining, and immunohistochemical staining for α-SMA, collagen I, and tumor growth factor-ß1 (TGF-ß1) were performed. RESULTS: Octreotide at a concentration of >20 ng/mL significantly inhibited PSC activation and proliferation in vitro. Inflammatory infiltration was reduced in the octreotide group in vivo, and the expression levels of α-SMA, collagen I, and TGF-ß1 were also lower, with statistic significant difference or not. Masson's trichrome staining showed a decrease in collagen deposition with octreotide treatment. CONCLUSIONS: Octreotide effectively inhibits PSC activation and proliferation in vitro, but has a limited inhibitory effect on the development of pancreas graft fibrosis.

Adoptive transfusion of tolerance dendritic cells prolongs the survival of cardiac allograft: a systematic review of 44 basic studies in mice.

BACKGROUND AND OBJECTIVE: Tolerogenic DCs (Tol-DCs), a group of cells with imDC phenotype, can stably induce T cells low-reactivity and immune tolerance. We systematically reviewed the adoptive transfusion of Tol-DCs induced by different ways to prolong cardiac allograft survival and its possible mechanism. METHOD: MEDLINE (1966 to March 2011), EMbase (1980 to March 2011), and ISI (inception to March 2011) were searched for identification of relevant studies. We used allogeneic heart graft survival time as endpoint outcome to analyze the effect of adoptive transfusion of Tol-DC on cardiac allograft. By integrating studies' information, we summarized the mechanisms of Tol-DC in prolonging cardiac grafts. RESULTS: Four methods were used to induce Tol-DC in all of the 44 included studies including gene-modified, drug-intervened, cytokine-induced, and other-derived (liver-derived & spleen-derived) DCs. The results showed that all types of Tol-DC can effectively prolong graft survival, and the average extension of graft survival time for each group was as follows: 22.02 ± 21.9 days (3.2 folds to control group) in the gene modified group, 25.94 ± 16.9 days (4.3 folds) in the drug-intervened groups, 9.00 ± 8.13 days (1.9 folds) in the cytokine-induced group, and 10.69 ± 9.94 days (2.1 folds) in the other-derived group. The main mechanisms of Tol-DCs to prolong graft survival were as follows: (1) induceT-cell hyporeactivity (detected by MLR); (2) reduce the effect of cytotoxic lymphocyte (CTL); (3) promote Th2 differentiation; (4) induce Treg; (5) induce chimerism. CONCLUSION: For fully MHC mismatched allogeneic heart transplant recipients of inbred mouse, adoptive transfusion of Tol-DC, which can be gene-modified, drug-intervened, cytokine-induced, spleen-derived or liver-derived, can clearly prolong the survival of cardiac allograft or induce immune tolerance. Gene-modified and drug-induced Tol-DC can prolong graft survival most obviously. Having better reliability and stability than drug-induction, gene-modification is the best way to induce Tol-DCs at present. One-time intravenous infusion of 2 × 10(6) Tol-DC is a simple and feasible way to induce long-term graft survival. Multiple infusions will prolong it but increase the risk and cost. Adoptive transfusion of Tol-DC in conjunction with immunosuppressive agents may also prolong the graft survival time.

A modified splint tubing technique for heterotopic heart transplantation in mouse.

BACKGROUND: The complexity of surgical procedure in mouse heterotopic heart transplantation (HHT) has prevented its widespread use. The present study reported a modified technique - splint tubing technique (STT) based on cuff technique (CT). MATERIALS AND METHODS: C57BL/10 and BALB/c mice were performed in syngeneic and allogeneic HHT using STT and CT. The main improvement is that the recipient external jugular vein and common carotid artery were independently opened a mouth and inserted a cannula to avoid the difficult operations of sleeved and everted tube. Graft function was assessed by pulse palpation, echocardiography and histopathologic examination. RESULTS: Ten syngeneic and thirty allogeneic HHT using STT were performed with six graft losses. Ten allogeneic HHT using CT were carried out with two graft losses. Technically successful syngeneic grafts have survived to the pre-specified 30days endpoint with strong contraction. STT significantly shortened operation time compared with CT (32.33±4.21min vs 45.15±4.89min, P<0.05). No significant difference was observed in survival time between two methods. CONCLUSION: STT is easily learned. It reduces the operation difficulty and makes the operation possible for the beginner to master this skill within 1-2weeks. Shorter operation time leads higher operative success rate.

Rb1 postconditioning attenuates liver warm ischemia-reperfusion injury through ROS-NO-HIF pathway.

AIMS: Ginsenoside Rb1 could prevent ischemic neuronal death and focal cerebral ischemia, but its roles to liver warm I/R injury remain to be defined. We determined if Rb1 would attenuate warm I/R injury in mice. MAIN METHODS: Mice were divided into sham, I/R, Rb1+I/R (Rb1 postconditioning, 20mg/kg, i.p. after ischemia), sham+L-NAME, I/R+L-NAME, and Rb1+I/R+L-NAME groups using 60min of the liver median and left lateral lobes ischemia. Serum levels of alanine aminotransferase (ALT) were measured and morphology changes of livers were evaluated. Contents of nitric oxide (NO) and nitric oxide synthase (NOS), malondialdehye (MDA) and activity of superoxide dismutase (SOD) were measured. Expressions of Akt, p-Akt, iNOS, HIF-1alpha, tumor necrosis factor-a (TNF-α) and intercellular adhesion molecule-1 (ICAM-1) were also determined by western blot or immunohistochemistry. KEY FINDINGS: Rb1 postconditioning attenuated the dramatically functional and morphological injuries. The levels of ALT were significantly reduced in Rb1 group (p<0.05). Rb1 upregulated the concentrations of NO, iNOS in serum, iNOS, and activity of SOD in hepatic tissues (p<0.05), while it dramatically reduced the concentration of MDA (p<0.05). Protein expressions of p-Akt, iNOS and HIF-1alpha were markedly enhanced in Rb1 group. Protein and mRNA expressions of TNF-α and ICAM-1 were markedly suppressed by Rb1 (p<0.05). SIGNIFICANCE: We found that Rb1 postconditioning could protect liver from I/R injury by upregulating the content of NO and NOS, and also HIF-1alpha protein expression. These protective effects could be abolished by L-NAME. These findings suggested Rb1 may have the therapeutic potential through ROS-NO-HIF pathway for management of liver warm I/R injury.

The effect of MICA antigens on transplant outcomes: a systematic review.

BACKGROUND AND OBJECTIVE: Human major histocompatibility complex class I-related gene A (MICA) is reportedly associated with poor transplant outcomes and a high risk of acute and chronic rejection in solid organ transplantation. However, studies on these risks have found conflicting results. In order to identify areas in which additional research is needed, we have undertaken the first systematic review of evidence concerning the risk of anti-MICA antibodies in recipients' sera. METHODS: We searched MEDLINE, EMBASE, and the Cochrane Library for original reports of clinical studies involving detection of MICA abs in transplant recipients' sera which used survival rate, acute rejection, and/or chronic rejection as outcome measures. RevMan 5.0.15 was used to calculate relative risk (RR), odds ratios (ORs), and 95% confidence intervals (95%CIs). RESULTS: We found 18 relevant articles, with a total of 6,607 recipients. Follow-up duration ranged from 1 to 15 years. In studies with more than 2 years of follow-up, anti-MICA abs positive in kidney recipients' post-transplant sera was associated with a lower graft survival rate (4 years: RR = 2.04, 95%CI 1.30 to 3.22; 3 years: OR = 3.56, 95%CI 1.47 to 8.62; 2 years: RR = 2.17, 95%CI 1.09 to 4.31) and a higher acute rejection rate (RR = 1.92, 95%CI 1.27 to 2.91), but there was no clear association with chronic rejection. Similar conclusions could not be drawn for heart or liver transplantation due to possible confounding by anti-HLA abs and the small sample sizes of the available studies. CONCLUSION: Anti-MICA antibodies in recipients' sera may associated with poor graft survival rates and high risk of acute and chronic rejection in solid organ transplantation, but more rigorous studies are needed to confirm or refute this relationship. Current immunosuppressive therapy may fail to suppress the harmful effect of MICA antigens.

The effects of immunosuppressive drugs on CD4(+) CD25(+) regulatory T cells: a systematic review of clinical and basic research.

OBJECTIVE: To review the effects of different immunosuppressive drugs on proliferation and function of regulatory T cells (Tregs). METHODS: We searched MEDLINE, Embase (from inception to September 2009), and the Cochrane Library (Issue 4, 2009) for clinical and basic research about the effects of various immunosuppressive drugs on Tregs. Data were extracted and methodological quality was assessed by two independent reviewers. Outcome measures for clinical research included blood Tregs levels, acute rejection episodes, and graft function. Outcomes for basic research included percentage of Tregs proliferation, function, Tregs phenotype, and evidence for possible mechanisms. We analyzed data qualitatively. RESULTS: Forty-two studies, including 19 clinical trials and 23 basic studies, were included. The immunosuppressive drugs studied were calcineurin inhibitors (CNIs), Rapa, anti-metabolism drugs, IL-2 receptor-blocking antibodies, T-cell depleting antibodies, and co-stimulation blockade antibodies. Most of the studies were on Rapa and CNIs. Eight basic studies on Rapa and CNIs showed that Rapa could promote the proliferation and function of Tregs, while CNIs could not. Five clinical trials involving a total of 158 patients showed that patients taking Rapa had higher blood concentration of Tregs than patients taking CNIs, but no difference was found in graft function (6-42 months follow-up). CONCLUSION: There is substantial evidence that Rapa favors Tregs survival and function. However, the higher numbers of blood Tregs in patients treated with Rapa do not show any association with better graft function. Larger clinical studies with longer follow-up are needed to more thoroughly assess the efficacy of immunosuppressive drugs on Tregs, and reveal whether a relationship exists between Tregs and graft function.

Hypoxia/reoxygenation up-regulates death receptor expression and enhances apoptosis in human biliary epithelial cells.

AIMS: To investigate whether ischemia/reperfusion (I/R)-induced apoptosis in the bile duct epithelium could be mediated by tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptors in biliary epithelial cells, we examined the effects of hypoxia/reoxygenation (H/R) on TRAIL cytotoxicity. MAIN METHODS: Using an H/R model, normal primary human intrahepatic biliary epithelial cells were exposed to hypoxia for 1 h, and then reoxygenated. Expressions of death receptor 4 (DR4) and DR5 mRNA and protein were measured. After 1 h of hypoxia, biliary epithelial cells were treated with TRAIL in different concentrations for 4 h. The death of biliary epithelial cells was confirmed by analysis of apoptosis and methylthiazolyl tetrazolium. The activities of caspase-3 and caspase-8 were determined by fluorometric assay. KEY FINDINGS: Compared with normoxic-cultured cells, the mRNA expressions of DR4 and DR5 were up-regulated from 0 min after reoxygenation, reaching a peak value at 60 min after reoxygenation. The protein expression of DR4 was most intense at 90 min after reoxygenation; the most intense expression of DR5 came at 120 min after reoxygenation. The apoptosis rate increased in the TRAIL treatment group and further increased in the TRAIL plus H/R group, and the effect of concentration-dependent TRAIL-mediated cell killing was more pronounced. Caspase-3 and caspase-8 enzymatic activities after H/R also increased with increased TRAIL concentration. SIGNIFICANCE: H/R up-regulated the expression of DR4 and DR5, and enhanced TRAIL-mediated apoptosis in normal human intrahepatic biliary epithelial cells.