Yanfei Huang

Murine Antithymocyte Globulin Therapy Alters Disease Progression in NOD Mice by a Time-Dependent Induction of Immunoregulation

OBJECTIVE—Antilymphocyte serum can reverse overt type 1 diabetes in NOD mice; yet, the therapeutic parameters and immunological mechanisms underlying the ability for this agent to modulate autoimmune responses against β-cells are unclear, forming the rationale for this investigation. RESEARCH DESIGN AND METHODS—A form of antilymphocyte serum, rabbit anti-mouse thymocyte globulin (mATG), was utilized in a variety of in vivo and in vitro settings, each for the purpose of defining the physiological, immunological, and metabolic activities of this agent, with particular focus on actions influencing development of type 1 diabetes. RESULTS—We observed that mATG attenuates type 1 diabetes development in an age-dependent fashion, only proving efficacious at disease onset or in the late pre-diabetic phase (12 weeks of age). When provided at 12 weeks of age, mATG reversed pancreatic insulitis, improved metabolic responses to glucose challenge, and rapidly increased frequency of antigen-presenting cells in spleen and pancreatic lymph nodes. Surprisingly, mATG therapy dramatically increased, in an age-dependent fashion, the frequency and the functional activity of CD4+CD25+ regulatory T-cells. Adoptive transfer/cotransfer studies of type 1 diabetes also support the concept that mATG treatment induces a stable and transferable immunomodulatory repertoire in vivo. CONCLUSIONS—These findings indicate that an induction of immunoregulation, rather than simple lymphocyte depletion, contributes to the therapeutic efficacy of antithymocyte globulin and suggest that time-dependent windows for the ability to delay or reverse type 1 diabetes exist based on the capacity to enhance the functional activity of regulatory T-cells.

Blocking Fas Ligand on Leukocytes Attenuates Kidney Ischemia-Reperfusion Injury

Inflammation contributes to the pathogenesis of ischemic acute kidney injury (AKI), and T cells mediate the early phase of ischemia-reperfusion injury (IRI). The Fas/Fas ligand (FasL) pathway modulates the balance of T cell subsets in the peripheral circulation as well as multiple inflammatory responses, suggesting that FasL may mediate ischemic AKI. Here, we induced bilateral renal IRI in mice bearing a loss-of-function mutation of FasL (the gld mutation) and in wild-type mice. Compared with wild-type mice, serum creatinine was lower in gld mice (1.4 ± 0.9 mg/dl versus 2.6 ± 0.4) at 24 hours after IRI (P<0.05). In addition, gld mice had fewer TNF-α-producing T lymphocytes in the kidneys and renal lymph nodes. Furthermore, pharmacologic blockade of FasL protected the kidneys of wild-type mice from IRI. Analysis of bone marrow chimeric mice suggested that the pathogenic effect of FasL involves leukocytes; reconstitution of wild-type mice with gld splenocytes attenuated IRI. In contrast, reconstitution of gld mice with wild-type splenocytes enhanced IRI. These data demonstrate that FasL, particularly on leukocytes, mediates ischemic AKI.

Kidney-Derived Stromal Cells Modulate Dendritic and T Cell Responses

Multipotent mesenchymal stromal cells from the bone marrow ameliorate acute kidney injury through a mechanism other than transdifferentiation into renal tissue. Stromal cells exert immunoregulatory effects on dendritic and T cells, both of which are important in the pathophysiology of immune-mediated kidney injury. We hypothesized that similar cells with immunoregulatory function exist within the adult kidney. We isolated murine kidney-derived cells with morphologic features, growth properties, and an immunophenotype characteristic of mesenchymal stromal cells. These cells lacked lineage markers and could be differentiated into mesodermal cell lineages, including osteocytes and adipocytes. Furthermore, these kidney-derived cells induced the generation of bone marrow-derived dendritic cells with significantly reduced MHC II expression, increased CD80 expression, increased IL-10 production and the inability to stimulate CD4+ T cell proliferation in allogeneic and nominal antigen-specific cultures. Experiments in mixed and transwell cultures demonstrated that the production of soluble immune modulators, such as IL-6, was responsible for these effects on dendritic cell differentiation and maturation. Contact-dependent mechanisms, however, inhibited mitogenic T cell proliferation. In summary, kidney-derived cells may suppress inflammation in the kidney in vivo; a better understanding of their biology could have therapeutic implications in a wide variety of immune-mediated kidney diseases.

Dendritic cell deficiency associated with development of BK viremia and nephropathy in renal transplant recipients

Background. BK virus nephropathy (BKVN) is a significant cause of renal allograft loss. Although overall intensity of immunosuppression is the greatest risk factor, recipient immune factors likely also play a role in the pathogenesis. Dendritic cells (DC) are potent antigen-presenting cells important for the induction of anti-viral cytotoxic T-cell responses. In a previous univariate analysis, we demonstrated a peripheral blood DC (PBDC) deficiency in patients with biopsy-proven BKVN, raising the possibility that reduction in DC predisposed to BK reactivation. Methods. In this study, we refined our previous analysis by comparing random posttransplant PBDC levels between an expanded group of patients with BKVN and controls without viremia using a multivariate analysis that accounted for factors known to influence PBDC levels. Next, we compared pretransplant PBDC levels between patients stratified by the presence or absence of posttransplant viremia. Finally, we assessed the predictive value of pretransplant PBDC levels for the development of posttransplant viremia. Results. Analyses revealed a PBDC level deficiency not only posttransplant in patients with BKVN but also pretransplant in patients who subsequently developed posttransplant BK viremia. Furthermore, we identified a pretransplant PBDC level that is a reasonable predictor for the development of posttransplant viremia. Conclusions. Our results identify PBDC deficiency as a previously unrecognized risk factor for BKV reactivation after renal transplantation. Pretransplant PBDC monitoring may prove to be a useful clinical tool in the assessment of patient vulnerability to BKVN posttransplant, which may allow more focused screening.

Kidney-derived mesenchymal stromal cells modulate dendritic cell function to suppress alloimmune responses and delay allograft rejection

Background. Mesenchymal stromal cells (MSCs) are multipotent cells with immunoregulatory capacity that are present in most adult organs. We previously demonstrated that co-culture of C57BL/6 kidney-derived MSCs (KSCs) in syngeneic bone marrow-derived dendritic cell (DC) culture induced a DC phenotype (KSC-DC) with reduced major histocompatibility complex (MHC) class II/increased CD80 expression and ability to suppress T-cell responses. Methods. To study their effects on allogeneic DCs, C57BL/6 KSCs were added to incipient BALB/c DC culture, with surface expression of MHC class II/CD80 measured by fluorescence-activated cell sorting. The ability to stimulate T-cell responses was then assessed in an allogeneic mixed leukocyte response. Next, we isolated either BALB/c (donor) or C57BL/6 (recipient) KSC-DCs from co-culture and measured the tempo of rejection after cotransplantation with islet grafts in a mouse model of islet transplantation. Finally, we measured the effects of KSC-DC stimulation on B-cell proliferation and IgM/IgG production in allogeneic cultures. Results. C57BL/6 KSCs induced a BALB/c DC phenotype with significantly decreased MHC class II, increased CD80 expression, and decreased T-cell stimulatory capacity in the mixed leukocyte response (P<0.01 vs. control). Cotransplantation of donor (BALB/c) but not recipient (C57BL/6) KSC-DCs resulted in significant delay of rejection after islet transplantation (P<0.01 vs. control). Finally, stimulation by KSC-DCs resulted in significantly reduced B-cell proliferation and antibody production in allogeneic culture (P<0.01 vs. control). Conclusions. Our results highlight an important mechanism of MSC-based immunotherapy and its potential for use in clinical transplantation as prevention of rejection and possibly sensitization.

Rabbit Polyclonal Mouse Antithymocyte Globulin Administration Alters Dendritic Cell Profile and Function in NOD Mice to Suppress Diabetogenic Responses

Mouse antithymocyte globulin (mATG) prevents, as well as reverses, type 1 diabetes in NOD mice, through mechanisms involving modulation of the immunoregulatory activities of T lymphocytes. Dendritic cells (DC) play a pivotal role in the generation of T cell responses, including those relevant to the autoreactive T cells enabling type 1 diabetes. As Abs against DC are likely generated during production of mATG, we examined the impact of this preparation on the phenotype and function of DC to elucidate novel mechanisms underlying its beneficial activities. In vivo, mATG treatment transiently induced the trafficking of mature CD8− predominant DC into the pancreatic lymph node of NOD mice. Splenic DC from mATG-treated mice also exhibited a more mature phenotype characterized by reduced CD8 expression and increased IL-10 production. The resultant DC possessed a potent capacity to induce Th2 responses when cultured ex vivo with diabetogenic CD4+ T cells obtained from BDC2.5 TCR transgenic mice. Cotransfer of these Th2-deviated CD4+ T cells with splenic cells from newly diabetic NOD mice into NOD.RAG−/− mice significantly delayed the onset of diabetes. These studies suggest the alteration of DC profile and function by mATG may skew the Th1/Th2 balance in vivo and through such actions, represent an additional novel mechanism by which this agent provides its beneficial activities.

Diverse Sources of Stem Cells to Treat Acute Kidney Injury

precursor cells (NPCs) can be protective in a rat ischemic AKI model. Fetal rat telencephalon-derived neurospheres were administered intravenously (i.v.), intraperitoneally (i.p.), and intracortically 4 h following 1 h of kidney ischemia. Control cells were homogenates from adult rat cerebellum. Intravenous administration of the neural precursor cells was highly effective in reducing the kidney dysfunction and structural damage at 24 h, and some benefit was seen from i.p. and intracortical administration as well. Labeled NPCs tracked to injured kidney, and there were enhanced levels of anti-inflammatory cytokines IL-4 and IL-10, while reduced levels of proinflammatory cytokines IL-1 and TNF in the kidney. Furthermore, macrophage infiltration was reduced after NPC administration. The authors are likely the first to demonstrate that NPCs are effective in AKI, which is an important conceptual advance in the field of stem cells and kidney diseases. A strength is that the design is clinically geared, studying various forms of administration. The injured kidney is providing homing signals that attract these progenitor cells, though what these are is unknown. The reduced macrophage trafficking and alterations in soluble inflammatory molecules are suggestive of an immune/anti-inflammatory mechanism of action, however reduced injury through other mechanisms may have led to this inflammatory profile. These fascinating results raise many questions. What are these NPCs making and could glial-derived neurotropic factor (GDNF) be one of these protective factors, as already demonstrated with GDNF administration [6, 7] ? Can other areas of the brain Ischemic acute kidney injury (AKI) decreases allograft survival in kidney transplants and increases mortality when occurring in native kidneys [1] . There is currently no specific treatment, thus there is an important need to develop novel ways to decrease early injury and augment repair. There is great promise in the application of stem cells for AKI, and the mechanisms by which stem cells work are largely unknown [2, 3] . Putative mechanisms of stem cell action have included transdifferentiation, fusion of stem cells into damaged tissue, immunomodulation, production of paracrine growth factors, or a combination of these mechanisms. Furthermore, it was once believed that stem cells originating from a particular tissue would support and construct tissue of similar type only. Recent studies challenge the notion of linear and unidirectional differentiation of lineage-committed adult stem cells. New lines of evidence invoke transdifferentiation to a wider spectrum of differentiated progeny than previously claimed [4, 5] . Undifferentiated or already differentiated cells or tissue-specific stem cells have been shown to alter their phenotype to express functional characteristics of a different tissue. These types of conversions are referred to as transdifferentiation or metaplasia and are accompanied by change in tissue-specific cell surface markers and phenotype. Another novel approach is to harness paracrine factors produced by stem cells and apply them to injured tissue to simulate stem cell repair function. In the current issue of Nephron, Wang et al. [pp e20– e28] from Brazil have elegantly demonstrated that neural Published online: April 3, 2009