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Abstract

Exposing donor mice to carbon monoxide (CO) protects transplanted islet allografts from immune rejection after transplantation (referred as the “donor” effect). In an attempt to understand the mechanisms of the donor effect of CO, we found that donor treatment with CO upregulates expression of peroxisome proliferatoractivated receptor γ (PPARγ), a transcriptional regulator, in isolated islets. In this study, we evaluated whether PPARγ contributes to the survival and function of transplanted islets and whether PPARγ mediates the protective effect of CO in a major mismatch islet allogeneic transplantation model. BALB/c (H-2^d) islets in which PPARγ activity was induced by its agonists, 15-deoxy-Δ^12–14-prostaglandin J₂ (15d-PGJ₂) or troglitazone were transplanted into C57BL/6 (H-2^b) recipients that had been rendered diabetic by streptozotocin (STZ). Blood glucose levels of recipients were monitored to determine the function of transplanted islets. Our data indicated that PPARγ activation in islets led to a high percentage of BALB/c islets survived long-term in C57BL/6 recipients. Activation of PPARγ in the donor suppresses expressions of proinflammatory cytokines including tumor necrosis factor-α (TNF-α) and inducible nitric oxide synthase (iNOS) in transplanted islets. Blocking PPARγ activity by its antagonist, GW9662, abrogated the donor effect of CO in vivo and in vitro. Our data demonstrate that PPARγ plays a critical role in the survival and function of transplanted islets after transplantation in the recipient. The protective effects of CO are at least in part mediated by PPARγ.

Keywords

Peroxisome proliferator-activated receptor γ (PPARγ)Islet transplantation Allograft survival

Introduction

Islet transplantation, which offers perhaps the greatest potential solution for the treatment of type 1 diabetes (T1D), involves immunosuppression of the recipient to prevent the immune system from rejecting transplanted grafts (46). Our recent findings provided the basis for an additional and complementary approach to the treatment of T1D: preconditioning of the donor. We found that induction of a protective gene, heme oxygenase-1 (HO-1), in or exposing its enzymatic product, carbon monoxide (CO), to islet donors not only resulted in long-term (>100 days) survival of transplanted islets in a highly significant percentage of allogeneic mouse recipients but also led to antigen-specific tolerance to transplanted islets (49). Ex vivo culturing of islets in a CO-saturated medium (CO at 1% was bubbled through the medium for 10 min) before transplantation also showed protection. This is important as a novel and powerful clinically relevant treatment modality for the treatment of T1D and other diseases.

Peroxisome proliferator-activated receptor γ (PPARγ) is a nuclear receptor involved in the control of various aspects of lipid metabolism (34). Upon activation, PPARγ heterodimerizes with retinoid X receptor, recruits specific cofactors, and binds to sequence-specific PPAR response elements in the promoter region of target genes. In this manner, PPARγ regulates several metabolic pathways, including lipid biosynthesis and glucose metabolism (38). PPARγ expression has been detected in kidney, liver, skeletal and smooth muscle, pancreas, small intestine, adipose tissue, and macrophages among others (2,13,14,16,30). Accumulating data show that PPARγ may play a role in modulation of the inflammatory response: activation of PPARγ results in a decrease in inducible nitric oxide synthase (iNOS) expression in mesangial cells, Kupffer cells, chondrocytes, and macrophages via its effect on nuclear factor-κ light-chain-enhancer of activated B cells (NF-κB), early growth response factor-1 (Egr-1), activator protein-1 (AP-1), and signal transducers and activators of transcription (STAT) pathways (12,15,31,41,47). PPARγ can form a transcriptional inhibitory complex with NF-κB and suppress the inducible expression of a wide variety of cellular and viral genes (8,17). Both PPARγ mRNA and protein are found in pancreatic β-cells (11). Dysfunction of PPARγ is related to diabetes, cardiovascular disease, obesity, and other diseases (24,29).

PPARγ can be activated by its natural and synthetic ligands. The most described natural ligands of PPARγ include lipoxygenase products [e.g., 13-hydroxy octadecadienoic acid (13-HODE) and 15-HETE] and cyclooxygenase products [prostaglandins (PGs), e.g. PGJ₂ and its derivative, 15d-PGJ₂] (19,36). Synthetic PPARγ agonists (e.g., troglitazone and rosiglitazone) act as insulin sensitizers and are currently used in the treatment of type 2 diabetes in clinic (5). In addition to diabetes, PPARγ agonists demonstrated therapeutic effects in animal models of ischemia/reperfusion, inflammation, shock, and others (25,37,51). They are also being successfully used in the treatment of a large range of human disorders including asthma (45), chronic obstructive pulmonary disease (18), cancers (27,28), pulmonary fibrosis (4), multiple sclerosis (40,43), and many other diseases (26).

In an effort to explain the donor effect of CO in the islet transplantation model, we observed that CO exposure to islet donors led to upregulated expression of PPARγ in isolated islets. Studies in macrophages also indicate that CO exerts its anti-inflammatory effects in part by induction of PPARγ, which otherwise would lead to an inflammatory response (3). Based on these facts, we evaluated whether CO treatment of the donors lead to increased PPARγ and therein offer a mechanism as to the survival benefits afforded by CO.

Materials and Methods

Animals

C57BL/6 and BALB/c mice at 6–8 weeks of age were purchased from the Jackson Laboratory (Bar Harbor, ME). The animal protocol was approved by the Animal Care Committee of the Beth Israel Deaconess Medical Center.

Islet Isolation and Transplantation

Donor mice were exposed to air or CO (250 ppm) in gas chambers for 1 h before islet harvesting. Islets were isolated as described elsewhere (22). Recipient mice were rendered diabetic by STZ (225 mg/kg IP, Sigma-Aldrich, St. Louis, MO). Five days after STZ administration, mice with two consecutive blood glucose levels exceeding 350 mg/dl were used as recipients. Islets (500–600 islet equivalent number, IEQ) were transplanted under the kidney capsule of the recipients. Blood glucose levels of the recipients were measured twice weekly with a glucometer (Roche, Basel, Switzerland) after transplantation. Mice with a blood glucose of < 200 mg/dl were considered normoglycemic. Grafts were deemed rejected when two consecutive glucose levels were > 300 mg/dl after a period of primary graft function.

Administration of PPARγ Agonists and Antagonist

Troglitazone and 15-d PGJ₂ were used to activate PPARγ, and 2-chloro-5-nitro-N-phenylbenzamide (GW9662) was used to block PPARγ activity in the study. 15-d PGJ₂ (0.4 mg/kg), troglitazone (1 mg/kg), and GW9662 (1 mg/kg) dissolved in 2% dimethyl sulfoxide (DMSO) were administered to the mice under anesthesia via penis vein injection. All chemicals were purchased from EMD Biosciences (Gibbstown, NJ) unless otherwise specified.

Real-Time RT-PCR Analysis

Total RNA was extracted using Qiagene RNA kit (Qiagen, Inc., Charworth, CA). DNase treatment was performed according to the manufacturer's suggestion (Qiagen) to prevent the contamination by genomic DNA. Real-time RT-PCR was performed to quantify the amount of target gene in each sample at mRNA level using the ABI PRISM® 7700 Sequence Detection Systems as described (49). Expressions of tumor necrosis factor (TNF)-α and iNOS were analyzed, and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) gene expression was quantified in each sample and used as endogenous control. 6-Carboxyfluorescein (FAM)-labeled real-time RT-PCR primers for TNF-α, iNOS, and GAPDH were purchased from Applied Bio systems (Life Technologies, Carlsbad, CA).

Western Blot

Cells or freshly harvested islets were washed in PBS, lysed in sodium dodecyl sulfate (SDS) sample buffer (2% SDS, 0.15 mol/L Tris, 10% mercaptoethanol, 10% glycerol, and 2 mmol/L phenylmethylsulfonyl fluoride), and separated in a 4–12% BT NuPAGE gel (Invitrogen, Carlsbad, CA). Proteins were transferred to a Hybond-P membrane (GE Healthcare, Piscataway, NJ) and blocked with 5% milk at room temperature for 60 min, followed by incubation with anti-PPARγ, anti-β-actin, or anti-IκBα antibodies (Santa Cruz Biotech). Blots were probed with corresponding secondary horseradish peroxidase antibodies (Cell Signal, Pierce, CA) and visualized by an ECL detection kit (Amersham Pharmacia Biotech, Little Chalfont, UK). The intensity of each signal was determined by the ImageJ software (NIH).

Apoptosis Assay and Detection of IκBα Degradation

Mouse insulinoma (βTC3) cells were seeded into six-well plates at a concentration of 1 × 10⁶ cells/well. Cells were pretreated with GW9662 (2 μM in 0.1% DMSO) or vehicle (0.1% DMSO) for 30 min before exposure to CO at 250 ppm in an Oxycycler (Biospherix, Lacona, NY). Cell death induced by recombinant murine interleukin-1β (IL-1β) (100 U/ml) and rat interferon-γ (IFN-γ) (1000 U/ml) (R&D Systems, Minneapolis, MN) was quantified by flow cytometry using the PE Annexin V Apoptosis Detection Kit (BD Pharmingen^™, Franklin Lakes, NJ) at 48 h after cytokine stimulation.

In a separate set of experiments, βTC3 cells stimulated with IL-1β and IFN-γ were collected at 0, 15, 30, and 60 min after cytokine stimulation. IκBα expression in whole-cell lysate was detected by Western blot using the anti-IκBα antibody. Expression of β-actin in each sample was measured and used as loading control.

Statistical Analyses

Kaplan–Meier survival curves were performed by using the Statview software, and the statistical differences were assessed by the log-rank test. Values of p < 0.05 were considered significant. Survival data are expressed as mean survival time±standard deviation (MST±SD). Differences between cytokine expressions were compared for statistical significance by the Student's t test with the Bonferroni correction.

Results

CO Administration to Donor Mice Upregulates PPARγ Expression in Isolated Islets

Islets were harvested from BALB/c donors exposed to CO (250 ppm, 1 or 4 h) or air (control) before islet isolation. PPARγ expression in freshly isolated, handpicked islets was evaluated by Western blot. Control islets harvested from air-treated donors expressed very low amounts of PPARγ (Fig. 1). In contrast, PPARγ expression was significantly increased in islets harvested from donors that had been exposed to CO for 1 or 4 h before isolation as compared to control (Fig. 1).

Figure 1.

CO exposure to BALB/c mice upregulated PPARγ expression in isolated islets. (A) Islets were isolated from mice exposed to carbon monoxide (CO) at 250 ppm (for 1 or 4 h) or air, and expressions of peroxisome proliferator-activated receptor γ (PPARγ) and β-actin were measured in freshly isolated islets by Western blot. (B) Relative expression of PPARγ as compared to β-actin level in each sample was quantitated by ImageJ software. Islets from CO-treated donors (dark gray and black bars) showed significantly increased expression of PPARγ at protein level as compared to islets harvested from air-treated donors (gray bar). Data shown are representative of at least three individual experiments. ∗p < 0.05 versus control as analyzed by Student's t test.

PPARγ Activation in Donor Mice Leads to Long-term Survival of Transplanted Islets in the Recipient in a Major Mismatch Islet Transplantation Model

To evaluate whether PPARγ activation in islets contributes to the survival and function of those islets in the recipient, PPARγ activity was induced in BALB/c donors by its agonists, 15d-PGJ₂ (0.4 mg/kg IV) and troglitazone (1 mg/kg IV), or blocked by its antagonist, GW9662 (1 mg/kg IV), 1 h before islet isolation. Donors in the control group received vehicle only (2% DMSO). Islets were transplanted into STZ-induced diabetic C57BL/6 recipients, and blood glucose levels of recipients were monitored. No further treatment was given to recipients. Islets from vehicle-treated donors were rejected in 17.8 ± 4.4 days (n = 5) when transplanted into untreated diabetic recipients. Blocking PPARγ activity with GW9662 in the donor led to graft rejection at 18.2 ± 3.5 days (n = 5), which was not significantly different when compared to control islets (p = 1.2 vs. control as analyzed by log-rank test). In contrast, induction of PPARγ by 15d-PGJ₂ led to three of four (75%) grafts surviving long-term (>100 days) in the recipients. Similarly, administering troglitazone to donor mice led to 50% long-term survival of islet allografts (n = 4) in recipients (Fig. 2). These data demonstrated that PPARγ activation in islets led to better survival and function of transplanted islets in a major mismatch islet transplantation model without any immunosuppression.

Figure 2.

Survival of BALB/c islets in which PPARγ was induced or suppressed in C57BL/6 recipients. BALB/c islets from donors that received vehicle [2% of dimethyl sulfoxide (DMSO)] alone were rejected in 17.8 ± 4.4 days when transplanted into C57BL/6 recipients that had been rendered diabetic by streptozotocin (STZ) (○, n = 5). Induction of PPARγ in the donors with 15-deoxy-prostaglandin J₂ (15d-PGJ₂) (■, n = 4) or troglitazone (▲, n = 4) led to 75% and 50% of islets survived long term in the recipient, respectively. Islets in which PPARγ was blocked by GW9662 (●, n = 5) were rejected as promptly as control islets.

PPARγ Activation Suppresses Expressions of TNF-α and iNOS in Transplanted Islets After Transplantation

Inflammation induced by stresses during islet isolation and transplantation contributes to apoptosis and rejection of transplanted islets/β-cells in the recipient. To assess whether PPARγ induction in the donor suppresses inflammation that occurs in transplanted islets, we measured expression of TNF-α and iNOS mRNA by real-time PCR in islet allografts at 1 day posttransplantation. Islet allografts from vehicle-treated donors had a dramatic increase in TNF-α and iNOS expression at 1 day posttransplantation as compared to freshly isolated islets (Fig. 3). In contrast, islets in which PPARγ activity was induced by 15d-PGJ₂ showed significantly less TNF-α and iNOS expression as compared to control (Fig. 3). Blocking PPARγ activity by GW9662 boosted TNF-α and iNOS in islet allografts, suggesting that PPARγ suppresses inflammation in transplanted islets that might contribute to the survival and function of those islets in the recipient.

Figure 3.

PPARγ activation suppressed expressions of TNF-α and iNOS in transplanted grafts posttransplantation. Islet allografts from PPARγ-induced donors (with 15d-PGJ₂, black bars) showed much less expressions of tumor necrosis factor-α (TNF-α) (A) and induced nitric oxide synthase (iNOS) (B) at 1 day posttransplantation as compared to grafts from vehicle-treated donors (light gray bars) or grafts from GW-treated donors (dark gray bars). ∗p < 0.025 versus control as analyzed by the Student's t test with the Bonferroni correction. Values were means of three to four grafts per group.

Blocking PPARγ Activity Abrogates the Effect of CO on Trans planted Islet Survival

To evaluate whether PPARγ mediates the donor effect of CO in the islet transplantation model, PPARγ activity was blocked by GW9662 (1 mg/kg IV, 1 h before) in BALB/c donors before CO exposure (250 ppm, for 1 h). Islets were then harvested and transplanted into diabetic C57BL/6 recipients. Blood glucose levels were monitored to determine the survival and function of transplanted islets. Mice that received vehicle (2% DMSO) before air exposure were used as controls. The mean graft survival time of control islets was 19.8 ± 5.0 days (Fig. 4). CO exposure to donors significantly prolonged islet survival with a mean survival time of 26.3 ± 6.7 days, with two grafts surviving >63 days at which time the experiment was terminated (p = 0.03 vs. control). In contrast, islets harvested from donors in which PPARγ activity was blocked were rejected promptly even in the presence of CO with a mean survival time of 20.6 ± 5.5 days (p = 0.15 vs. control). Our data thus indicate that PPARγ activity is required for CO to impart benefit when treating donors in the islet allogeneic transplantation model.

Figure 4.

Blocking PPARγ activity by GW9662 abrogates the donor effect of CO. BALB/c islets from control donors were rejected in 19.8 ± 5.0 days in C57BL/6 recipients (●, n = 5). CO exposure to donor mice prolonged islet survival to 26.3 ± 6.7 days, with two grafts survived >63 days at the time when the experiment was terminated (▲, n = 6, p = 0.03 vs. control). Blocking PPARγ activity with GW before CO exposure reduced graft survival to 20.6 ± 5.5 days, which was not significantly different from islets from control donors (○, n = 5, p = 0.15 vs. control).

CO Protects βTC3 Cells From Cytokine-Induced Apoptosis In Vitro in a PPARγ-Dependent Manner

We have shown that CO exposure protected an insulinoma cell line, βTC3 cells, from cytokine-induced apoptosis (23). We tested here whether PPARγ activity is essential for the antiapoptotic effect of CO in β-cells. Cells that had been pretreated with GW9662 (2 μM, 30 min before CO exposure) or vehicle (0.1% DMSO) were exposed to CO at 250 ppm for 2 h before being stimulated with IL-1β (100 U/ml) plus IFN-γ (1,000 U/ml). Cell apoptosis was quantitated 48 h after cytokine treatment by flow cytometry using Annexin-V and 7-aminoactinomycin D (7-AAD) staining. Cytokine treatment led to 27.9 ± 5.3% of cells undergoing apoptosis. Preexposure of cells to CO before cytokine stimulation reduced cell death to 10.3 ± 2.1%, which was completely lost in cells in which PPARγ activity was blocked (29.3 ± 5.3% of cells underwent apoptosis, which was not significantly different from control). Nonstimulated cells exhibit 2.4 ± 0.6% cell death. These results demonstrate that PPARγ activity is necessary for the antiapoptotic effect of CO in β-cells.

CO protects βTC3 cells from cytokine-induced apoptosis via preventing degradation of IκBα, the inhibitor of NF-κB (Wang et al., unpublished data). We tested whether PPARγ is essential for CO to inhibit IκBα degradation/ NF-κB activation. PPARγ activity was blocked in βTC3 cells by GW9662 before CO exposure and cytokine stimulation as described above. Expression of IκBα in cells at 0, 15, 30, and 60 min after cytokine administration was analyzed by Western blot. IL-β and IFN-γ led to a rapid degradation of IκBα in βTC3 cells; that is, IκBα was completely absent at 15 min after cytokine treatment and its expression then returned to normal 60 min later (Fig. 5A). In contrast, significantly less IκBα degradation was observed in cells preexposed to CO (250 ppm, 2 h) before stimulation with cytokines. Again, CO failed to block IκBα degradation when PPARγ activity was inhibited by GW9662, that is, IκBα was completed degraded in groups pre-treated with GW9662 before CO exposure, even though elevated PPARγ expression was observed in cells exposed to CO (Fig. 5B). Again, our data confirmed that PPARγ activity is required for CO to suppress IκBα degradation/NF-κB activation in βTC3 cells.

Figure 5.

Blocking PPARγ with GW9662 in βTC3 abrogates the antiapoptotic effect of CO in vitro. (A) Exposing mouse insulinoma (βTC3) cells to interleukin (IL)-1β and interferon (IFN)-γ led to 27.9 ± 5.3% of the cells undergoing apoptosis. Preexposure cells to CO before cytokine-stimulation reduced cell death to 10.3 ± 2.1% of the cells. In contrast, cells in which PPARγ was blocked by GW9662 before CO exposure had 29.3 ± 5.3% of the cells undergo apoptosis, which is not different from the control group. Nonstimulated cells have 2.4 ± 0.6% of cell death. (B) CO failed to inhibit IκBα degradation when PPARγ activity was blocked. Cytokine exposure to βTC3 cells led to a rapid degradation of IκBα at 15 min after cytokine administration. CO exposure prevented IκBα degradation (top), and IκBα was completely degraded when PPARγ activity was blocked by GW9662 even with CO exposure (bottom).

Discussion

The most significant finding of our study is that activation of PPARγ in the donor leads to a high percentage of transplanted islets survived long-term in the recipient without immunosuppression. Additionally, that donor treatment with inhaled CO induces PPARγ in islets, which prolongs islet cell survival. Similar protective effects of CO have also been observed in macrophages in an endotoxic shock model (3).

While the results of human islet transplantation are promising, this technique is still in the experimental stage (42,44). Major hurdles that hinder its clinical application are apoptosis of transplanted islets caused by nonimmune mediated factors and the toxicity of immunosuppressive drugs used to suppress the immune rejection response mounted against transplanted islets by the recipient. Our data indicate that donor treatment leads to long-term survival and tolerance of transplanted islets in recipients in the absence of immunosuppressants in a murine islet allogeneic transplantation model (49).

We induced PPARγ activity by two different PPARγ agonists, 15d-PGJ₂, a natural ligand of PPARγ, and troglitazone, a high-affinity synthetic PPARγ ligand. Both treatments dramatically enhanced islet allograft survival in a major mismatch islet transplantation model. These treatments are as effective or even more dramatic than that observed with CO exposure alone. As PPARγ agonists have been approved for clinical use as insulin sensitizers to treat patients with type 2 diabetes and other diseases, they can potentially be prescribed for islet transplantation in the treatment of T1D based on data presented here.

Accumulating evidence indicate that inflammation plays a major role in the rejection of transplanted islets/ β-cells after transplantation and tolerance cannot be achieved in the presence of inflammation (7,22,33,35). In our previous studies, we showed that the donor effect of CO was primarily due to reduction of inflammation normally present in islets following transplantation. CO, when given to the donor, suppresses the expression of proinflammatory cytokines [e.g., TNF-α, iNOS, monocyte chemotactic protein-1 (MCP-1), etc.] that likely contribute to graft rejection through induction of apoptosis. To evaluate whether induction of PPARγ has a similar anti-inflammatory effect as that observed with CO, we measured expression of TNF-α and iNOS in islet allografts from PPARγ-treated donors 1 day posttransplantation. As expected, islets in which PPARγ was induced showed much less TNF-α and iNOS expression as compared to controls. In contrast, inhibition of PPARγ activity using the selective inhibitor GW9662 boosted TNF-α and iNOS expression. It thus seems that the anti-inflammatory property of PPARγ plays a principal role in contributing to islet allograft survival and function after transplantation.

We conclude that PPARγ agonists exert their protection via two major mechanisms. First, PPARγ agonists protect β-cells from apoptosis. As observed in studies from others and us, early β-cell death induced by stresses elicited during islet harvesting and transplantation process represents a major obstacle for islet survival after transplantation. Even under optimal conditions, approximately 60% of islets die within 3 days after transplantation due to nonimmune-mediated factors in a syngeneic islet transplantation model (9). Thus, preventing early β-cell death, we view as critical to prolong graft survival. We observed that induction of PPARγ protects β-cells from cytokine-induced apoptosis. Evidence is also accumulating in the literature that PPARγ induction by its agonists promotes function of β-cells via modulating endoplasmic reticulum (ER) stress pathways (21), protecting against islet amyloid polypeptide and lipo-toxicities (50), and reducing oxidative stress (6,48). Thus, treatment with PPARγ agonists inhibits nonimmune-mediated β-cell death that might contribute to their survival and function. Secondly, we provide evidence that PPARγ agonists suppress inflammation in transplanted islets induced by proinflammatory cytokines, reactive oxygen species, and other stresses. This may be caused by a reduced cell infiltration around islet allografts after transplantation. However, the exact mechanism by which PPARγ activation leads to graft acceptance is still under investigation. We suspect that its anti-inflammatory property plays a major role in this process. According to the danger hypothesis (32), an immune response in the presence of danger signals such as inflammatory cytokines (20) becomes uncontrolled, whereas in the absence of danger, tolerance ensues (1,39). As tolerance can only be achieved in a less inflammatory environment, PPARγ activation reduced inflammation and led to graft acceptance in the recipient even in the absence of immunosuppression.

As described in our other study evaluating the anti-inflammatory effect of CO in a model of acute lung injury in mice, the induction of PPARγ in islets was most likely transient, that is, its expression level might well decrease after CO exposure ends as it is only expressed as a transcription factor to regulate downstream gene activation such as Egr-1, a central mediator of inflammation (3).

Based on similar donor effects of CO exposure and induction of PPARγ in response to donor exposure to CO, we hypothesized that PPARγ activity is required for the protective effects of CO. To test this hypothesis, we blocked PPARγ activity with GW9662, before CO exposure in both in vivo and in vitro models. In the in vivo model, we compared survival time of islet allografts harvested from donor mice in which PPARγ was blocked prior to CO exposure. In the in vitro model, we measured cytokine-induced cell apoptosis and degradation of IκBα in βTC3 cells. In both experimental settings, blocking PPARγ activity abrogated the protective effect of CO, which correlated that PPARγ activity is required for the protective effects of CO.

Considering that human islets after transplantation into patients with T1D will be challenged with an auto immune response within the recipient, it would be extremely beneficial if PPARγ induction can protect islet grafts from autoimmune rejection. Based on the dramatic effect of PPARγ activation, the ability to modulate PPARγ activity is certainly an approach that can be readily applied in the clinic to improve islet graft survival and function in both the autologous and allogeneic islet transplantation settings (10). These possibilities are currently being investigated.

In conclusion, induction of PPARγ in donor mice using PPARγ agonists or inhaled CO prolongs survival and function of transplanted islets in an allogeneic islet transplantation model. Understanding the protective effects of CO and PPARγ will help develop treatment regimens to treat and hopefully cure diabetes and other diseases.

Footnotes

Acknowledgments

This work was supported in part by JDRF grants 5-2006-911, 5-2007-989, and 1-2007-619 to H.W. The authors declare no conflicts of interest.

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Activation of Peroxisome Proliferator-Activated Receptor γ Prolongs Islet Allograft Survival

Abstract

Keywords

Introduction

Materials and Methods

Animals

Islet Isolation and Transplantation

Administration of PPARγ Agonists and Antagonist

Real-Time RT-PCR Analysis

Western Blot

Apoptosis Assay and Detection of IκBα Degradation

Statistical Analyses

Results

CO Administration to Donor Mice Upregulates PPARγ Expression in Isolated Islets

PPARγ Activation in Donor Mice Leads to Long-term Survival of Transplanted Islets in the Recipient in a Major Mismatch Islet Transplantation Model

PPARγ Activation Suppresses Expressions of TNF-α and iNOS in Transplanted Islets After Transplantation

Blocking PPARγ Activity Abrogates the Effect of CO on Trans planted Islet Survival

CO Protects βTC3 Cells From Cytokine-Induced Apoptosis In Vitro in a PPARγ-Dependent Manner

Discussion

Footnotes

Acknowledgments

References