IL-10 Gene Transfer Research Articles

Gene Ex Vivo Transduction Effects on Flap Survival and MicroRNAs Expression Analysis in Rat Allotransplantation Model

The lack of noninvasive biomarkers for graft rejection remains a challenge for the accurate monitoring of vascularized composite allotransplants. Viral vector-mediated gene transfer is a promising method for preventing graft rejection. In this study, we aimed to establish the expression profile of microRNAs (miRNAs) in skin flap allotransplantation, with or without gene transfer, and determine the potential role of several miRNAs as biomarkers of acute rejection and immune tolerance. An abdominal epigastric flap was transplanted from SD (RT1a) to Wistar rats (RT1Au). The adenoviral interleukin 10 (vIL-10) gene was transferred to the experimental group via flap pedicle injection. Postoperatively, flap appearance, hematoxylin and eosin staining, immunohistochemical staining, and miRNA expression analyses were performed. The viral IL-10 gene-treated group showed improved flap survival and reduced acute rejection response compared with the control group. On postoperative day 7, IL-10 expression in the flap was identified using immunohistochemistry and real-time polymerase chain reaction. The expression of miR-191a, miR-31a, miR-16, and miR-3473 was upregulated in the skin tissue, and that of miR-484, miR-132, miR-139, miR-150, and miR-6216 was upregulated in the serum. AV IL-10 gene transfer could be an effective immunosuppressive strategy for the prevention of skin flap allograft rejection. Additionally, some miRNAs were upregulated in the experimental group, serving as potential biomarkers of immune tolerance.

Eggshell membrane modulates gut microbiota to prevent murine pre-cachexia through suppression of T helper cell differentiation.

BackgroundCachexia is a life‐threatening condition observed in several pathologies, such as cancer or chronic diseases. Interleukin 10 (Il10) gene transfer is known to improve cachexia by downregulating Il6. Here, we used an IL10‐knockout mouse model to simulate cachexia and investigate the effects of eggshell membrane (ESM), a resistant protein, on general pre‐cachexia symptoms, which is particularly important for the development of cachexia therapeutics.MethodsFive‐week‐old male C57BL6/J mice were fed an AIN‐93G powdered diet (WT), and 5‐week‐old male B6.129P2‐Il10 < tm1Cgn>/J (IL10−/−) mice were fed either the AIN‐93G diet (KO) or an 8% ESM‐containing diet (KOE) for 28 weeks. The tissue weight and levels of anaemia‐, blood glucose‐, lipid metabolism‐, and muscular and colonic inflammation‐related biochemical markers were measured. Transcriptomic analysis on liver and colon mucus and proteomic analysis on skeletal muscle were performed. Ingenuity Pathway Analysis was used to identify molecular pathways and networks. Caecal short‐chain fatty acids (SCFAs) were identified using HPLC, and caecal bacteria DNA were subjected to metagenomic analysis. Flow cytometry analysis was performed to measure the CD4+ IL17+ T cells in mesenteric lymph nodes.ResultsThe body weight, weight of gastrocnemius muscle and fat tissues, colon weight/length ratio, plasma HDL and NEFA, muscular PECAM‐1 levels (P < 0.01), plasma glucose and colonic mucosal myeloperoxidase activity (P < 0.05) and T helper (Th) 17 cell abundance (P = 0.071) were improved in KOE mice over KO mice. Proteomic analysis indicated the protective role of ESM in muscle weakness and maintenance of muscle formation (>1.5‐fold). Transcriptomic analysis revealed that ESM supplementation suppressed the LPS/IL1‐mediated inhibition of RXR function pathway in the liver and downregulated the colonic mucosal expression of chemokines and Th cell differentiation‐related markers (P < 0.01) by suppressing the upstream BATF pathway. Analysis of the intestinal microenvironment revealed that ESM supplementation ameliorated the microbial alpha diversity and the abundance of microbiota associated with the degree of inflammation (P < 0.05) and increased the level of total organic acids, particularly of SCFAs such as butyrate (2.3‐fold), which could inhibit Th1 and Th17 production.ConclusionsESM supplementation ameliorated the chief symptoms of cachexia, including anorexia, lean fat tissue mass, skeletal muscle wasting and reduced physical function. ESM also improved colon and skeletal muscle inflammation, lipid metabolism and microbial dysbiosis. These results along with the suppressed differentiation of Th cells could be associated with the beneficial intestinal microenvironment and, subsequently, attenuation of pre‐cachexia. Our findings provide insights into the potential of ESM in complementary interventions for pre‐cachexia prevention.

Adeno-Associated Viral Vector–Mediated Interleukin 10 Gene Transfer in Sprague-Dawley Rat Skin Allograft

Viral vector-mediated interleukin 10 gene transfer is one of the most promising methods for the prevention of graft rejection. Adeno-associated virus (AAV) is a nonpathogenic helper-dependent parvovirus. This is one of the most promising vehicles for gene delivery. Our study aimed to analyze the immune-suppressive potential of recombinant adeno-associated virus-mediated rat IL-10 in rat skin allograft. Dorsal skin from Sprague-Dawley rats was grafted to Fischer 344 rats. In vivo AAV-viral IL-10 (vIL-10) gene transfer was done in the experimental group by direct subcutaneous injection. Observation of graft appearance, cytologic and immunohistochemical testing, and confocal immunofluorescence were performed at postoperative days 5 and 10. The viral IL-10 gene-treated group showed improved graft survival and reduced acute rejection response compared to the control group. IL-10 expression on skin was identified by immunohistochemistry and confocal microscopy. AAV IL-10 gene transfer could be an effective immunosuppressive method for preventing skin allograft rejection.

Generation of Powerful Human Tolerogenic Dendritic Cells by Lentiviral-Mediated IL-10 Gene Transfer.

The prominent role of dendritic cells (DC) in promoting tolerance and the development of methods to generate clinical grade products allowed the clinical application of tolerogenic DC (tolDC)-based therapies for controlling unwanted immune responses. We established an efficient method to generate tolerogenic human DC, producing supra-physiological levels of IL-10, by genetically engineering monocyte-derived DC with a bidirectional Lentiviral Vector (bdLV) encoding for IL-10 and a marker gene. DCIL−10 are mature DC, modulate T cell responses, promote T regulatory cells, and are phenotypically and functionally stable upon stimulation. Adoptive transfer of human DCIL−10 in a humanized mouse model dampens allogeneic T cell recall responses, while murine DCIL−10 delays acute graft-vs.-host disease in mice. Our report outlines an efficient method to transduce human myeloid cells with large-size LV and shows that stable over-expression of IL-10 generates an effective cell product for future clinical applications in the contest of allogeneic transplantation.

Viral Vector Mediated Interleukin-10 Gene Transfer in Skin Allograft

BackgroundExpression of genes with immunoregulatory capacity can potentially decrease rejection of allograft. According to recent studies, viral interleukin (IL)-10 can reduce immune response during allotransplantation and is one of the most promising methods for the prevention of rejection. Our study aimed to analyze the immunosuppressive potential of recombinant adenovirus-mediated rat IL-10 in rat skin allograft. MethodsWe performed skin graft surgery 1 hour after infecting the donated skin with adenovirus-mediated rat IL-10. On day 7 postoperatively, the skin allografts were harvested, and acute rejection was graded histologically. ResultsViral IL-10 gene transfer into rat skin allografts improved graft survival and reduced acute rejections. ConclusionThe results of our study suggest that the therapeutic potential of graft viral IL-10 gene transfer is an effective immunosuppressive method for preventing skin allograft rejection.

Engineered T Regulatory Type 1 Cells for Clinical Application.

T regulatory cells, a specialized subset of T cells, are key players in modulating antigen (Ag)-specific immune responses in vivo. Inducible T regulatory type 1 (Tr1) cells are characterized by the co-expression of CD49b and lymphocyte-activation gene 3 (LAG-3) and the ability to secrete IL-10, TGF-β, and granzyme (Gz) B, in the absence of IL-4 and IL-17. The chief mechanisms by which Tr1 cells control immune responses are secretion of IL-10 and TGF-β and killing of myeloid cells via GzB. Tr1 cells, first described in peripheral blood of patients who developed tolerance after HLA-mismatched fetal liver hematopoietic stem cell transplantation, have been proven to modulate inflammatory and effector T cell responses in several immune-mediated diseases. The possibility to generate and expand Tr1 cells in vitro in an Ag-specific manner has led to their clinical use as cell therapy in patients. Clinical grade protocols to generate or to enrich and expand Tr1 cell medicinal products have been established. Proof-of-concept clinical trials with Tr1 cell products have demonstrated the safety and the feasibility of this approach and indicated some clinical benefit. In the present review, we provide an overview on protocols established to induce/expand Tr1 cells in vitro for clinical application and on results obtained in Tr1 cell-based clinical trials. Moreover, we will discuss a recently developed protocol to efficient convert human CD4+ T cells into a homogeneous population of Tr1-like cells by lentiviral vector-mediated IL-10 gene transfer.

Efficacy of In Vivo Electroporation-Mediated IL-10 Gene Delivery on Survival of Skin Flaps.

Despite advances in understanding the underlying mechanisms of flap necrosis and improvement in surgical techniques, skin flap necrosis after reconstructive surgery remains a crucial issue. We investigated the efficacy of electroporation-mediated IL-10 gene transfer to random skin flap with an aim to accelerate wound healing and improve skin flap survival. Nine male Wistar rats (300-330g) were divided in two groups (a) control group (n=5), only surgery no gene transfer, and (b) experimental group, received electroporation-mediated IL-10 gene transfer 24h before the surgery as prophylaxis (n=4). Random skin flap (McFarlane) was performed in both groups. Planimetry, Laser Doppler imaging, and immunohistochemistry were used to evaluate the effect of IL-10 gene transfer between study groups at day 7. Electroporation-mediated IL-10 gene transfer decreased percentage of flap necrosis (p value=0.0159) and increased cutaneous perfusion compared to the control group (p value=0.0159). In addition, Spearman's rank correlation showed a significant negative correlation between percentage of flap necrosis and Laser Index (p value=0.0083, r -0.83, respectively). Furthermore, significantly higher mean CD31+ vessel density was detected in the experimental group compared to the control group (p value=0.0159). Additionally, semi-quantitative image analysis showed lower inflammatory cell count in experimental group compared to control group (p value=0.0317). In vivo electroporation-mediated IL-10 gene transfer reduced necrosis, enhanced survival and vascularity in the ischemic skin flap.

Hydrodynamic IL10 Gene Transfer in Human Colon: Results from an "EX VIVO" Study with Potential Clinical Application in Crohn's Disease.

The aim of this work is to evaluate the efficacy of hydrodynamic venous IL10 gene delivery to "ex vivo" human colon segments and to determine its potential interest in Crohn's disease treatment. Twenty human colon segments were obtained from surgical resections. Hydrodynamic transfection through the main vein of the pedicle with 50 mL of hIL10 plasmid (20 μg/mL) solution was performed on 13 of them. Tissue sections were cultured and DNA, RNA, and protein copies were determined after 1, 2, and 4 days. Data obtained were compared with 6 nontransfected specimens. Finally, 1 specimen was injected with gold nanoparticles, and their distribution was examined under electron microscope. IL10 DNA levels were higher in treated tissues than in controls (P < 0.001), decreasing along time. The amount of hIL10 RNA was significantly increased in treated tissues when compared with controls (P = 0.001). The indexes of protein IL10 translation in treated groups were much higher (P < 0.001) than the basal production. The protein expression was higher in transfected tissue (10-50-fold, with respect to control tissue); this difference being established during the first hours and maintained during, at least, 4 days. With electron microscopy, we hardly observed large (15 nm) gold nanoparticles within the tissue, always in the submucosa. However, multiple small (4 nm) nanoparticles were observed within the cytoplasm of enterocytes in mucosa. Hydrodynamic procedure efficiently delivers the IL10 gene to the human colon, achieving levels of tissue protein expression high enough to mediate pharmacological effects with interest in controlling immune response in patients with Crohn's disease.

Mesenchymal Stem Cells Overexpressing Interleukin-10 Promote Neuroprotection in Experimental Acute Ischemic Stroke

Interleukin (IL)-10 is a contributing factor to neuroprotection of mesenchymal stem cell (MSC) transplantation after ischemic stroke. Our aim was to increase therapeutic effects by combining MSCs and ex vivo IL-10 gene transfer with an adeno-associated virus (AAV) vector using a rat transient middle cerebral artery occlusion (MCAO) model. Sprague-Dawley rats underwent 90 min MCAO followed by intravenous administration of MSCs alone or IL-10 gene-transferred MSCs (MSC/IL-10) at 0 or 3 hr after ischemia reperfusion. Infarct lesions, neurological deficits, and immunological analyses were performed within 7 days after MCAO. 0-hr transplantation of MSCs alone and MSC/IL-10 significantly reduced infarct volumes and improved motor function. Conversely, 3-hr transplantation of MSC/IL-10, but not MSCs alone, significantly reduced infarct volumes (p < 0.01) and improved motor function (p < 0.01) compared with vehicle groups at 72 hr and 7 days after MCAO. Immunological analysis showed that MSC/IL-10 transplantation significantly inhibits microglial activation and pro-inflammatory cytokine expression compared with MSCs alone. Moreover, overexpressing IL-10 suppressed neuronal degeneration and improved survival of engrafted MSCs in the ischemic hemisphere. These results suggest that overexpressing IL-10 enhances the neuroprotective effects of MSC transplantation by anti-inflammatory modulation and thereby supports neuronal survival during the acute ischemic phase.

The influence of AAV2-mediated gene transfer of human IL-10 on neurodegeneration and immune response in a murine model of Parkinson's disease

BackgroundThe aim of this study was to examine the effect of AAV2-hIL-10 (vector containing cDNA for human interleukin 10) on dopaminergic system activity (measured as DA levels and TH mRNA expression in mouse striata), and other monoamine and amino acid neurotransmitters concentration as well as development of inflammatory processes (measured as TGF-β, IFN-γ and GFAP mRNA expression) in a murine MPTP neurotoxicant model of Parkinson's disease. MethodsMale C57BL/6 mice 12 months-old were used in this study. AAV2-hIL-10 vector was bilaterally administered into striatum at 14, 21 or 28 days prior to MPTP intoxication. Animals were sacrificed at 7 days following MPTP injection. The expression of hIL-10 (human interleukin 10) was examined by ELISA. Striatal monoamine and amino acid neurotransmitters were measured by HPLC method. TH, TGF-β, IFN-γ and GFAP mRNA expression was examined by RT-PCR method. ResultsMPTP treatment dramatically reduced DA levels and decreased TH mRNA expression in mouse striata, effects that were significantly impeded by AAV2-hIL-10 administration prior to MPTP intoxication. AAV2-hIL-10 infusion increased IFN-γ, TGF-β and GFAP mRNA expression. ConclusionsOur data suggest that the transfer of AAV2-hIL-10 into the striatum may play a neuroprotective role in the mouse MPTP model of PD and these effects are mediated by the anti-inflammatory action of IL-10.

T-Cell-Mediated Suppression: From Bench to Bedside

T-Cell-Mediated Suppression: From Bench to Bedside

Farelerde İntraabdominal Sepsis Modelinde İntravenöz ve İntraperitoneal İnterlökin 10 Lipozom Aracılı Gen Tedavisinin Karşılaştırılması

The most novel approach utilizing IL-10 in sepsis is IL-10 gene delivery in experimental model of sepsis. In our study, we aimed to compare kinetics of intravenous versus intraperitoneal delivery of IL-10 gene transfer in early stages of sepsis. This is a prospective controlled experimental study. Six groups were gathered with 20 Swiss-Albino female mice. Intra-abdominal sepsis was induced by cecal ligation and puncture (CLP). Animals had either intraperitoneal or intravenous IL-10 liposomal gene transfer. Animals were sacrified 24 h after injections, followed by harvest of lung,liver,spleen,vena cava tissues. Immunostaining revealed more prominent staining in liver after intraperitoneal delivery. All endothelial tissues stained with intravenous delivery. There was striking difference between tissue expressions of transgene of animals in CLP intravenous group when compared to other groups. Our results point out that pro-inflammatory action of IL-10 is prominent in intravenous gene delivery which shows itself with induction of zyon. IL-10 still may harbor therapeutical potential which still needs to be explored.

The intensity of bronchiolar epithelial cell injury caused by an alloimmune response is ameliorated by transbronchial human interleukin-10 gene transfer in a rat model of lung transplantation

The aim of the present study was to determine whether the intensity of bronchiolar epithelial cell injury is ameliorated by transbronchial human IL-10 (hIL-10) gene transfer in a rat model of lung allograft rejection. The left lung was extracted from a donor Brown Norway rat and transbronchially instilled with encoding hIL-10 (IL-10 group) or β-galactosidase (control group). The lung graft was then transplanted into a Lewis rat and harvested on day 6 after transplantation. The allografts were histologically examined and all bronchioles in the slides were assigned one of three grades according to the intensity of epithelial cell loss. The distribution of the grades was significantly different between the two groups, and the epithelial cell injury was significantly improved in the IL-10 group. The present study demonstrated the effect of transbronchial hIL-10 gene transfer on ameliorating bronchiolar epithelial cell injury in a rat model of lung allograft rejection.

Early gestational gene transfer of IL-10 by systemic administration of lentiviral vector can prevent arthritis in a murine model

Gene therapy has been applied to murine models of rheumatoid arthritis (RA) using a number of different strategies to downregulate inflammation in synovial joints. However, prolonged joint expression has been problematic. Our laboratory has found that early gestational intravascular injection of lentiviral vector leads to efficient transduction and sustained transgene expression in articular cartilage and synovium. In this study, we show that in utero gene transfer of IL-10 can prevent and decrease pathology in a murine model of RA. Following prenatal injection of lentiviral vector containing murine IL-10 gene, the cytokine was detectable in the serum, and the green fluorescent protein reporter gene was detectable in chondrocytes and synoviocytes of adult mice up to 21 weeks of age. Adult mice that had been treated prenatally were later immunized against type II collagen to induce an autoimmune arthritis. Compared with controls, prenatally treated mice demonstrated delayed onset of arthritis, decreased frequency of arthritis and markedly decreased severity of disease, by both clinical and histological criteria. This effect was directly related to levels of IL-10 expression, but no immunosuppressive effects of the therapy were observed. This study demonstrates proof of principle for the prenatal prevention and amelioration of RA by early gestational gene transfer of the anti-inflammatory cytokine, IL-10.

Early gestational gene transfer of IL-10 by lentiviral vector can prevent arthritis in a murine model

Early gestational gene transfer of IL-10 by lentiviral vector can prevent arthritis in a murine model

Effects of adenovirus-mediated interleukin-10 gene transfer on apoptosis and insulin secretion function of beta cell

To evaluate the efficacy of adenovirus vector-mediated murine interleukin-10 (mIL-10) gene transfer to rat beta cell-RINm5F cells in vitro and to explore the potential value of gene therapy in type 1 diabetes mellitus. The recombinant adenovirus vector Ad-mIL-10 was constructed and transfected into RINm5F cells (Ad-mIL-10 group). Untransfected RINm5F cells and Ad-eGFP-transfected cells were used as controls. The expression of mIL-10 was examined by RT-PCR and Western blot. The levels of IL-10 in supernatant were measured by enzyme-linked immunosorbent assay (ELISA). For a determination of insulin release, the cells were cultured with high glucose (16.7 mmol/L) for a 1-hour co-incubation. Then radioimmunoassay was used to detect the level of insulin in supernatant. After induction of IL-1beta, the levels of nitric oxide (NO) and nitric oxide synthase (NOS) were measured, the apoptosis of transfected cells was detected by Hoechst 33258 staining and Fas expression by flow cytometry. Both mRNA and protein of Ad-mIL-10 were successfully expressed in RINm5F cells. Expression of mIL-10 gene resulted in significant increases in insulin secretion in response to high glucose. Compared with uninfected control and Ad-eGFP infected group, Ad-mIL-10 infected group had decreased levels of NO and NOS induced by IL-1beta [NO level (nmol/10(6) cells): 52.9 +/- 3.2 vs. 227.3 +/- 26.4, 235.1 +/- 28.6, both P < 0.05; NOS level (U/10(6) cells): 9.3 +/- 1.2 vs. 29.8 +/- 2.5, 30.5 +/- 2.8, both P < 0.05]. Furthermore, Ad-mIL-10 gene transfer led to a profound reduction of Fas-expressing islet cells under the induction of IL-1beta. Fas-expressing islet cells in Ad-mIL-10 group were significantly lower than those in uninfected group and Ad-eGFP-transfected group (24.6% +/- 1.0% vs. 33.3% +/- 5.1%, 32.6% +/- 1.1%) (P < 0.05). The apoptotic rates of Ad-mIL-10 group were lower in comparison with the other two groups (9.4% +/- 1.1% vs. 19.2% +/- 2.2%, 20.6% +/- 2.3%, P < 0.05). IL-10 gene transfer to islet beta cells may be beneficial in maintaining beta cells function, protecting islet cells from IL-1beta-mediated apoptosis and promoting islet cells survival. It is a potential therapy for type 1 diabetes mellitus.

Adeno-Associated Virus–Mediated IL-10 Gene Transfer Suppresses Lacrimal Gland Immunopathology in a Rabbit Model of Autoimmune Dacryoadenitis

To evaluate the effect of adeno-associated virus (AAV) vector-mediated viral (v)IL-10 gene expression on lacrimal gland (LG) immunopathology and ocular surface disease in a rabbit model of induced autoimmune dacryoadenitis (ID). Autologous peripheral blood lymphocytes, activated in a mixed-cell reaction when cocultured with purified rabbit lacrimal epithelial cells, induce a Sjögren's-like autoimmune dacryoadenitis when injected directly back into the donor animal's inferior LG. Four weeks after disease induction, AAV vector expressing the vIL-10 gene under control of a tetracycline-inducible promoter was injected into the inferior LG of the treatment group (ID/Rx), and doxycycline was fed orally to induce transgene expression. The ID group serving as control also received doxycycline. All LGs were removed 16 weeks after disease induction. Clinical symptoms showed overall improvement in the ID/Rx group compared with the ID group. Histopathologic examination of the ID group's LG revealed scattered large lymphocytic foci and areas of altered or distorted acini, whereas the ID/Rx group had scattered small lymphocytic foci. The number of CD18(+) cells was almost fivefold lower in the ID/Rx group than in the ID group. Although the total number of RTLA(+) cells did not differ between the groups, the CD4/CD8 ratio was 16-fold smaller in the ID/Rx group. Animals with experimentally induced autoimmune dacryoadenitis appeared to benefit from AAV-mediated vIL-10 gene transfer therapy. Quantitative immunohistochemical analysis suggested that the therapy might not have been simply immunosuppressive but rather supported the induction of CD8(+) regulatory cells.

344. Early Gestational Gene Transfer of IL-10 by Lentiviral Vector Can Prevent Arthritis in a Murine Model

344. Early Gestational Gene Transfer of IL-10 by Lentiviral Vector Can Prevent Arthritis in a Murine Model

β-tubulin-induced autoimmune hearing loss exacerbates in IL-10-deficient mice and restores by IL-10 gene transfer (143.10)

Abstract IL-10 is a potent immunoregulatory cytokine in homeostatic regulation of autoreactive T cell repertoire, experimental autoimmune hearing loss (EAHL) is characterized by Th1 cell-mediated disorder plus down-regulation of IL-10. We hypothesized that endogenous levels of IL-10 function to regulate the severity of EAHL, and exogenous of IL-10 would abrogate EAHL. BALB/c (WT) and homozygous IL-10-deficient mice were underwent β-tubulin immunization to develop EAHL; some β-tubulin-immunized IL-10-deficient mice were intramuscular injected IL-10 DNA. EAHL developed progressively in both WT and IL-10-deficient mice immunized with β-tubulin, and severity of hearing loss in IL-10-/- mice was significantly greater than that in WT animals. However, the hearing function was significantly improved in IL-10 DNA administration group than that in the control group, ABR thresholds are equivalent to naive controls, and histological analysis of cochlear cross section was similar to those of naïve controls. IL-10 DNA administration suppressed autoreactive T cells and reduced expression of IFN-γ, the proportion of regulatory T cells (Treg) of IL-10-treated mice was significantly increased in comparison with control mice. Therefore lack of IL-10 exacerbates hearing loss, exogenous administration of IL-10 improved hearing function. Thus, IL-10 is capable of controlling autoimmune reaction severity by suppressing Th1-type inflammatory production and enhancing the percentage of Treg cells.

Induction of Lymphocyte Apoptosis in Rat Liver Allograft by Adenoviral Gene Transfection of Human Interleukin-10

Background/Aims: Gene therapy can provide a possible avenue in organ transplantation to treat acute allograft rejection. This study was designed to investigate the effect of adenovirus-mediated human IL-10 (hIL-10) gene transfer on the apoptosis of infiltrating lymphocytes and examine the efficacy of hIL-10 gene transfer in combination with subtherapeutic doses of cyclosporine A (CsA) in a rat liver transplantation model. Methods: Inbred male DA and LEW rats were used for liver donors and recipients, respectively. The rats were divided into saline, Ad-lacZ, CsA, Ad-hIL-10 and Ad-hIL-10 + CsA groups. Graft survival, histopathological, enzyme-linked immunosorbent assay, reverse transcriptase-polymerase chain reaction and flow cytometry were performed in liver specimens obtained from different time points after transplantation in the 5 groups. Results: Ad-hIL-10 pretreatment inhibited allograft rejection, prolonged the survival of hepatic allografts, and downregulated the expression of IFN-γ and IL-2 mRNA, with simultaneous upregulation of IL-4 mRNA. In addition, Ad-hIL-10 pretreatment upregulated the expression of Fas mRNA in the isolated graft-infiltrating lymphocytes and induced graft-infiltrating lymphocyte apoptosis. A single subtherapeutic dose of CsA acted synergistically with it. Conclusion: hIL-10 gene therapy induced alloreactive lymphocyte apoptosis via Fas/FasL pathway. hIL-10 gene transfection in combination with subtherapeutic doses of CsA facilitates the long-term survival of liver grafts.