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R Zhong

Publications and source records attributed to R Zhong.

At least 19 recordsLinked to original sources
PubMed2007

Anti-CD45RB monoclonal antibody prolongs renal allograft survival in cynomolgus monkeys.

Previously, an anti-CD45RB monoclonal antibody (mAb) has been shown to induce murine allograft tolerance. The present study was performed to assess the ability of an anti-human CD45RB mAb to prevent rejection in a monkey MHC-mismatched kidney transplant model. The recipients were allocated into the following treatment groups: (1) isotype control IgG; (2) mouse anti-human CD45RB IgG1 (6G3); (3) human-mouse chimeric anti-CD45RB-IgG1 (C6G3-IgG1); (4) human-mouse chimeric anti-CD45RB-IgG2 (C6G3-IgG2); (5) tacrolimus at a subtherapeutic dose and (6) tacrolimus and C6G3-IgG1 in combination. Monotherapy with anti-CD45RB mAb significantly prolonged renal allograft survival to a median survival of 21 days. Adding a subtherapeutic dose of tacrolimus improved the efficacy of the anti-CD45RB mAb, achieving a median survival of 85 days, whereas a subtherapeutic dose of tacrolimus alone only moderately prolonged survival to 27 days. Treatment with anti-CD45RB mAb resulted in an alteration of the CD45RB(hi) : CD45RB(lo) cell ratio in the peripheral blood. We have, for the first time, demonstrated that an anti-human CD45RB mAb (6G3) can prolong graft survival. Induction with an anti-CD45RB mAb improves the efficacy of tacrolimus in the prevention of rejection. These encouraging results indicate that an anti-CD45RB mAb may be valuable in future clinical transplantation.

Animals↗
PubMed2007

Gal knockout and beyond.

Recently, Galalpha1-3Galbeta1-4GlcNAc (Gal) knockout (k/o) pigs have been developed using genetic cloning technologies. This remarkable achievement has generated great enthusiasm in xenotransplantation studies. This review summarizes the current status of nonhuman primate experiments using Gal k/o pig organs. Briefly, when Gal k/o pig organs are transplanted into primates, hyperacute rejection does not occur. Although graft survival has been prolonged up to a few months in some cases, the overall results were not better than those using Gal-positive pig organs with human complement regulatory protein transgenes. Gal k/o pig kidneys rapidly developed rejection which was associated with increased anti-non-Gal antibodies. Although the precise mechanisms of Gal k/o pig organ rejection are not clear, it could result from incomplete deletion of Gal, up-regulation of new antigen (non-Gal antigen) and/or production of non-Gal antibodies. Future work in xenotransplantation should place emphasis on further modification of donors, such as combining human complement regulatory genes with Gal k/o, deleting non-Gal antigens and adding protective/surviving genes or a gene that inhibits coagulation. Induction of donor-specific T- and B-cell tolerance and promotion of accommodation are also warranted.

Animals↗
PubMed2006

NOS2 (iNOS) deficiency in kidney donor accelerates allograft loss in a murine model.

Renal NOS2 is expressed and produces abundant nitric oxide (NO) in various renal cells in response to proinflammatory cytokines. However, the role of this enzyme in renal allograft survival remains unknown. Kidney allotransplantation was performed in the murine model of C57BL/6J (H-2(d)) to nephrectomized Balb/c (H-2(b)) mice. Here we show that deficiency in NOS2 expression in kidney donors significantly advanced allograft failure, indicated by decreasing mean survival of recipients receiving NOS2 null grafts (15.4 +/- 6.4 days) as compared to those with wild type grafts (65.4 +/- 28.1 days) (p = 0.0005). Consistent with survival results, NOS2 null grafts had more severe renal tubule injury and decreased renal function compared to wild type grafts. In vitro NOS2 expressing TEC had greater resistance to allogeneic lymphocyte-mediated apoptosis. The addition of exogenous NO inhibited Fas-mediated TEC apoptosis and reduced proliferation of allogeneic lymphocytes. These data suggest that endogenous production of NO through renal NOS2 activity can play a protective role in kidney grafts through attenuating Fas-mediated donor cell apoptosis as well as by inhibiting proliferation of inflammatory infiltrating lymphocytes. Enhanced donor NOS2 expression may be a useful strategy to improve kidney transplant survival.

Animals↗
PubMed2006

Prolongation of allograft survival by administration of anti-CD45RB monoclonal antibody is due to alteration of CD45RBhi: CD45RBlo T-cell proportions.

CD45RB monoclonal antibody (mAb) therapy is capable of prolonging allograft survival. We have previously shown that CD45RB mAb enriches the CD45RBlo T-cell population in vitro and in vivo by preferentially depleting CD45RBhi T cells. The present study assessed the importance of CD45RBhi T-cell depletion in murine cardiac allograft survival by infusion of naive CD45RB T-cell subsets. Here we show that naturally occurring CD45RBloCD4+ T cells express regulatory transcription factor Foxp3 and have regulatory function, whereas CD45RBhiCD4+ T cells express low levels of Foxp3 and have effector function. Infusion of syngeneic CD45RBhi T cells significantly reduced graft survival after depletion of CD45RBhi T cells by CD45RB mAb. Reduction of graft survival also occurred when syngeneic CD45RBhi T cells were infused into rapamycin-treated mice, whereas survival was prolonged when CD45RBlo T cells were added. This indicates that an alteration in the balance between regulatory CD45RBlo and effector CD45RBhi T cells is critical to the immunologic function of CD45RB mAb. A strategy to eliminate effector T cells with consequent enrichment of the regulatory T-cell compartment may be an important new strategy in the prevention of rejection.

Animals↗
PubMed2006

Preventing renal ischemia-reperfusion injury using small interfering RNA by targeting complement 3 gene.

The complement system is one of the important mediators of renal ischemia-reperfusion injury (IRI). We hypothesized that efficient silencing of C3, which is the central component on which all complement activation pathways converge, could be achieved using small interfering RNA (siRNA), and that this would result in overall inhibition of complement activation, thereby preventing IRI in kidneys. A series of experiments was conducted, using a mouse model of IRI and vector-delivered C3-specific siRNA. We demonstrated the following: (1) renal expression of C3 increases as a result of IRI; (2) by incorporation into a pRNAT U6.1 vector, siRNA can be delivered to renal cells in vivo; (3) systemically delivered siRNA is effective in reducing the expression of C3 in an experimentally induced mouse kidney model of IRI; (4) similarly, siRNA reduces complement-mediated IRI-related effects, both in terms of renal injury (as evidenced by renal function and histopathology examination) and mouse mortality and (5) silencing the production of C3 diminishes in vivo production of TNF-alpha. This study implies that siRNA represents a novel approach to preventing IRI in kidneys and might be used in a variety of clinical settings, including transplantation and acute tubular necrosis.

Animals↗
PubMed2006

A synergistic effect between PG490-88 and tacrolimus prolongs renal allograft survival in monkeys.

This study was undertaken to determine if PG490-88 and tacrolimus (Tac) act synergistically to prevent renal allograft rejection in monkeys and to explore possible mechanisms of synergy between these agents. MHC-mismatched renal allografts were transplanted into cynomolgus monkeys after bilateral nephrectomy. Recipients were divided into the following groups: (i) no treatment; (ii) PG490-88 (0.03 mg/kg); (iii) Tac (1 mg/kg); (iv) PG490-88 (0.01 mg/kg) + Tac (1 mg/kg) and (v) PG490-88 (0.03 mg/kg) + Tac (1 mg/kg). Through synergy PG490-88 and Tac inhibited anti-CD3/PMA-induced T-cell proliferation and IFN-gamma expression in vitro. Tac monotherapy only marginally prolonged survival (27 +/- 3.2 days), while the combination of PG490-88 and Tac significantly prolonged graft survival to a median of 99 days (PG490-88 at 0.03 mg) and 38.5 days (PG490-88 at 0.01 mg/kg). Prolonged survival correlated with inhibited IgM production as well as reduced T-cell infiltration, IL-2 protein expression and NF-AT/NF-kappaB activity. We conclude that PG490-88 and a subtherapeutic dose of Tac significantly prolong renal allograft survival in monkeys through the synergistic inhibition of T-cell activation and a decrease in IFN-gamma production and NF-AT/NF-kappaB activity.

Animals↗
PubMed2006

Increasing resistance of tubular epithelial cells to apoptosis by shRNA therapy ameliorates renal ischemia-reperfusion injury.

Renal tubular epithelial cells (TEC) die by apoptosis or necrosis in renal ischemia-reperfusion injury (IRI). Fas/Fas ligand-dependent fratricide is critical in TEC apoptosis, and Fas promotes renal IRI. Therefore, targeting Fas or caspase-8 may have therapeutic potential for renal injury in kidney transplant or failure. RNA silencing by short hairpin RNA (shRNA) is a novel strategy to down-regulate protein expression. Using this approach, silencing of Fas or caspase-8 by shRNA to prevent TEC apoptosis and IRI was evaluated. IRI was induced by renal artery clamping for 45 or 60 min at 32 degrees C in uninephrectomized C57BL/6 mice. Here, we showed that Fas or pro-caspase-8 expression was significantly knocked down in TEC by stable expression of shRNA, resulting in resistance to apoptosis induced by superoxide, IFN-gamma/TNF-alpha and anti-Fas antibody. Inferior vena cava delivery of pHEX-small interfering RNA targeting Fas or pro-caspase-8 resulted in protection of kidney from IRI, indicated by reduction of renal tubular injury (necrosis and apoptosis) and serum creatinine or blood urea nitrogen. Our data suggest that shRNA-based therapy targeting Fas and caspase-8 in renal cells can lead to protection of kidney from IRI. Attenuation of pro-apoptotic proteins using genetic manipulation strategies such as shRNA might represent a novel strategy to promote kidney allograft survival from rejection or failure.

Animals↗
PubMed2003

Renal tubular epithelial cell apoptosis by Fas-FasL-dependent self-injury can augment renal allograft injury.

The role of Fas-FasL interactions in kidney allograft injury may be complex as renal tubular epithelial cells (TEC) express both Fas and FasL. The role and regulation of TEC self-injury has not been investigated. In co-cultures of TEC, FasL-bearing, Fas-null TEC was demonstrated to induce apoptosis of TEC-bearing Fas. Co-culturing effector lpr-TEC (M3.1-lpr) with target WT-TEC (CS3.7) at a ratio of 10:1 (E/T) induced 15.2 +/- 2.4% of target apoptosis as compared to its basal level of 2.6 +/- 0.3%. Similarly lpr-TEC induced apoptosis in gld-TEC (MRM-gld) from a basal level of 3.7 +/- 0.2% to 6.4 +/- 0.3%. Expression of kidney Fas-FasL on injury was tested in a renal transplant model. C57BL/6 (B6) mice were transplanted with Fas-deficient C3H-lpr/lpr or FasL mutation C3H-gld/gld kidneys as compared to normal (wild-type [WT]) C3H/Hej donors. Survival of both lpr and gld recipient was improved compared to WT donors (P <.05) as was function of lpr and gld kidneys indicated by a lower serum creatinine (LPR: 41 +/- 8 micromol/L; GLD: 52 +/- 7 micromol/L) as compared to the WT donors (84 +/- 8 micromol/L, P <.001). These results demonstrate that activated TEC may commit a novel and previously unreported form of self-injury (fractricide) through Fas-FasL. These results suggest that inhibition of renal Fas or FasL might be a useful strategy to prevent TEC loss during rejection.

Animals↗
PubMed2002

Thymic re-entry of mature activated T cells and increased negative selection in vascularized allograft recipients.

Transplantation tolerance is a dynamic state that involves several homeostatic mechanisms intrinsic to the host. One of these mechanisms is activation-induced T cell death (AICD). However, it is unclear where AICD takes place during alloreactive responses. Since activated T cells can re-enter the thymus, we hypothesized that mature T cells activated by an allograft could be deleted upon re-entry into the thymus. To test this hypothesis, we used wild-type or 2C TCR transgenic mice receiving syngeneic or allogeneic heterotopic, vascularized heart grafts. First, we demonstrated that ex vivo CFSE-labelled T cells re-entered the thymus when transferred into allograft recipients but not when transferred into isograft recipients. Next, we compared the changes in cell subset numbers and incidence of apoptosis in the thymi and spleens of allograft or isograft recipients. Seven days after transplantation, at a time in which all the allografts were undergoing rejection, cells expressing donor-MHC class II molecules had migrated to the thymus and to the spleen. In the thymus of allograft recipients, overall cellularity was significantly reduced by 40% and associated with an increase in the number of double negative (CD4-CD8-) thymocytes and a decrease in double positive (CD4+CD8+) thymocytes, consistent with increased negative selection of thymocytes. Additionally, thymi of allograft recipients showed an increase in the number of recently activated, mature T cells (TCRhi, CD25+, CD44+) and a significant increase in the number of apoptotic cells, especially in the thymic medulla, that involved mature T cells as indicated by the TCRhi, CD44+, CD4 or CD8 single positive phenotype. Spleens of allograft recipients were increased in size and cellularity but did not show any of the changes in cell subsets seen in the thymi. Our data show that after allografting there is an increase in apoptotic cell death that is associated with negative selection of developing thymocytes as well as of alloreactive mature T cells that have re-entered the thymus upon activation in the periphery. This may occur upon migration of graft-derived antigen-presenting cells to the thymus.

Animals↗
PubMed2001

Spleen autotransplantation in mice: a novel experimental model for immunology study.

The aim of the experimental model to develop a spleen transplant model in mice to study the role of spleen in autoimmune and transplant rejection. After a midline incision, splenectomy was performed. Four tiny segments were cut from the removed spleen and were rinsed at room temperature in physiological salt solution. The greater omentum was lifted and four omental pockets were created; four thin segments were then placed into the "nests," subsequently marked, and fixed using 8-0 suture. The abdomen was then closed. The duration of the survival time was different among the nine groups (n = 3-3). Tissue samples were taken from the marked areas for histological examination stained with hematoxylin and eosin (H&E). H&E staining demonstrated large, well-circumscribed splenic nests with lymphoid zone and red pulp and well-formed trabecules in the spleen. Among the possible applications of this novel model is the ability to study the role of spleen in autoimmune and organ rejection.

Animals↗
PubMed2001

Fragmentation of N-oxides (deoxygenation) in atmospheric pressure ionization: investigation of the activation process.

The diagnostic fragmentation of N-oxides resulting from loss of the oxygen atom (MH+ --> MH+-O) in electrospray and atmospheric pressure chemical ionization (APCI) mass spectra was investigated. When the temperature of the heated capillary tube was elevated, the ratio of the intensity of the [MH+ -16] fragment to the precursor ion (MH+) increased. This 'deoxygenation' process was associated with thermal activation and did not result from collisional activation in the desolvation region of the API source. Although the extent of 'deoxygenation' is compound-dependent, it can provide evidence for the presence of an N-oxide in a sample and can be used to distinguish N-oxides from hydroxylated metabolites (Ramanathan et al. Anal. Chem. 2000; 72: 1352). To demonstrate the practical application of thermal fragmentation of N-oxides, liquid chromatography (LC)/APCI-MS was used to distinguish an N-oxide drug from its hydroxylated metabolite in an unprocessed rat urine sample, despite the fact that the drug and its metabolite were not fully resolved by HPLC.

Animals↗
PubMed2001

A short course of high-dose cyclophosphamide induces long-term survival of intestinal allografts in mice.

Several transplant programs have recently added cyclophosphamide (CyP) to their immune suppression protocols in an attempt to reduce intestinal graft rejection rates. The present study was undertaken to confirm the benefits of this drug in a murine small bowel transplant model. A short course of monotherapy with CyP 20 mg/kg per dose resulted in a mean survival time (MST) of 17.5 +/- 3.6 days, compared with a MST of 7.5 +/- 0.7 days in the untreated controls (P < 0.01). Cyclosporin A (CsA) 30 mg/kg per day produced comparable survival rates when used as monotherapy (MST: 14.2 +/- 1.3 days) or in combination with CyP 20 mg/kg per dose (MST: 21.3 +/- 5.1 days). Treatment with high dose CyP (40 mg/kg per dose) completely prevented graft loss in 8 of 10 animals (MST: 72.5 +/- 5.3 days, P < 0.01). However, adding CsA abrogated the induction of long-term survival achieved by CyP alone (MST: 23 +/- 0.4 days). These data have important implications for the use of CyP in clinical transplantation.

Animals↗
PubMed2001

Vascular smooth muscle cells of recipient origin mediate intimal expansion after aortic allotransplantation in mice.

Intimal expansion by vascular smooth muscle cells (SMCs) is a characteristic feature of graft vascular disease. Whether graft intimal SMCs arise from donor or recipient tissue is not well established but has important pathogenetic implications. We examined for the presence of male cells in the expanded intima of sex-mismatched mouse aortic allografts (C57BL/6-to-BALB/c) at 30 or 60 days after transplant by in situ hybridization using a Y-chromosome probe. Study groups included male-to-female allografts, female-to-male allografts, and female-to-female allografts in recipients previously engrafted with male bone marrow. Although intimal expansion developed in all allografts, male-to-female allografts lacked Y-chromosome-positive intimal cells. In contrast, such cells were abundant in female-to-male allografts and most of these cells co-labeled for smooth muscle alpha-actin by immunostain. Female-to-female allografts in recipients with male bone marrow showed a limited number of intimal Y-chromosome-positive cells. However, none of these clearly co-labeled for smooth muscle alpha-actin and their numbers declined throughout time, consistent with graft-infiltrating inflammatory cells. We conclude that intimal expansion of mouse aortic allografts is mediated by SMCs that originated from the recipient. There was little evidence of their derivation from the bone marrow, suggesting instead the adjacent host aorta as the primary source of intimal SMCs.

Animals↗
PubMed2001

Caffeoyl coenzyme A O-methyltransferase and lignin biosynthesis.

Lignin, a complex phenylpropanoid compound, is polymerized from the monolignols p-coumaryl alcohol, coniferyl alcohol and sinapyl alcohol. These three monolignols differ only by the 3- and 5-methoxyl groups. Therefore, enzymatic reactions controlling the methylations of the 3- and 5-hydroxyls of monolignol precursors are critical to determine the lignin composition. Recent biochemical and transgenic studies have indicated that the methylation pathways in monolignol biosynthesis are much more complicated than we have previously envisioned. It has been demonstrated that caffeoyl CoA O-methyltransferase plays an essential role in the synthesis of guaiacyl lignin units as well as in the supply of substrates for the synthesis of syringyl lignin units. Caffeic acid O-methyltransferase has been found to essentially control the biosynthesis of syringyl lignin units. These new findings have greatly enriched our knowledge on the methylation pathways in monolignol biosynthesis.

Lignin↗
PubMed2001

Alteration of auxin polar transport in the Arabidopsis ifl1 mutants.

The INTERFASCICULAR FIBERLESS/REVOLUTA (IFL1/REV) gene is essential for the normal differentiation of interfascicular fibers and secondary xylem in the inflorescence stems of Arabidopsis. It has been proposed that IFL1/REV influences auxin polar flow or the transduction of auxin signal, which is required for fiber and vascular differentiation. Assay of auxin polar transport showed that the ifl1 mutations dramatically reduced auxin polar flow along the inflorescence stems and in the hypocotyls. The null mutant allele ifl1-2 was accompanied by a significant decrease in the expression level of two putative auxin efflux carriers. The ifl1 mutants remained sensitive to auxin and an auxin transport inhibitor. The ifl1-2 mutant exhibited visible phenotypes associated with defects in auxin polar transport such as pin-like inflorescence, reduced numbers of cauline branches, reduced numbers of secondary rosette inflorescence, and dark green leaves with delayed senescence. The visible phenotypes displayed by the ifl1 mutants could be mimicked by treatment of wild-type plants with an auxin polar transport inhibitor. In addition, the auxin polar transport inhibitor altered the normal differentiation of interfascicular fibers in the inflorescence stems of wild-type Arabidopsis. Taken together, these results suggest a correlation between the reduced auxin polar transport and the alteration of cell differentiation and morphology in the ifl1 mutants.

Arabidopsis↗
PubMed2001

Anti-CD45RB monoclonal antibody-mediated transplantation tolerance.

Currently, lifelong immunosuppression is required for organ transplant recipients. The majority of transplant recipients will eventually develop chronic rejection with resultant graft loss, despite treatment with powerful immunosuppressive agents. These agents are also associated with numerous toxicities including reduced immunity against infection and malignancy. Therefore, the central goal in transplant science is to devise tolerance strategies in an attempt to establish a state of prolonged non-reactivity against the allograft, accompanied with preservation of an intact immune system. Although predictable tolerance induction has been elusive, we found that short course of the novel immunomodulatory agent, anti-CD45RB monoclonal antibody, leads to indefinite acceptance of renal allografts in mice, and has been shown to markedly prolong allograft survival in primates. We review the current state of development of this antibody, and the progress made in defining its mechanism of action.

Adoptive Transfer↗