Masayoshi Miura

Neutralization of Groα and Macrophage Inflammatory Protein-2 Attenuates Renal Ischemia/Reperfusion Injury

Previous studies have provided strong evidence for a role for neutrophils in mediating pathology during reperfusion of ischemic tissues. CXC chemokines including interleukin-8, KC/Groα, and macrophage inflammatory protein (MIP)-2, direct neutrophils to tissue sites of inflammation. In the current study we tested the efficacy of antibodies to KC/Groα and MIP-2 in inhibiting neutrophil infiltration into kidneys during reperfusion after 1 hour of warm ischemia using a mouse model. KC mRNA and protein were produced within 3 hours after reperfusion of the ischemic kidneys. MIP-2 mRNA and protein were twofold to fourfold lower than KC and were at low levels until 9 hours after reperfusion. Only 60% of mice subjected to ischemia/reperfusion injury survived to day 3 after reperfusion. Treatment with rabbit neutralizing antibodies to both KC and MIP-2 inhibited neutrophil infiltration into ischemic kidneys during reperfusion, restored renal function as assessed by decreased serum creatinine and urea nitrogen levels to near normal levels, and resulted in complete survival of treated animals. Finally, treatment with both antibodies significantly reduced histologically graded pathology of kidneys subjected to ischemia/reperfusion injury. Collectively, the results indicate the efficacy of neutralizing the chemokines directing neutrophils into ischemic kidneys during reperfusion to inhibit this infiltration and attenuate the resulting pathology.

Monokine Induced by IFN-γ Is a Dominant Factor Directing T Cells into Murine Cardiac Allografts During Acute Rejection

The use of chemokine antagonism as a strategy to inhibit leukocyte trafficking into inflammatory sites requires identification of the dominant chemokines mediating recruitment. The chemokine(s) directing T cells into cardiac allografts during acute rejection remain(s) unidentified. The role of the CXC chemokines IFN-γ inducible protein 10 (IP-10) and monokine induced by IFN-γ (Mig) in acute rejection of A/J (H-2a) cardiac grafts by C57BL/6 (H-2b) recipients was tested. Intra-allograft expression of Mig was observed at day 2 posttransplant and increased to the time of rejection at day 7 posttransplant. IP-10 mRNA and protein production were 2.5- to 8-fold lower than Mig. Whereas allografts were rejected at day 7–9 in control recipients, treatment with rabbit antiserum to Mig, but not to IP-10, prolonged allograft survival up to day 19 posttransplant. At day 7 posttransplant, allografts from Mig antiserum-treated recipients had marked reduction in T cell infiltration. At the time of rejection in Mig antiserum-treated recipients (i.e., days 17–19), intra-allograft expression of macrophage-inflammatory protein-1α, -1β, and their ligand CCR5 was high, whereas expression of CXCR3, the Mig receptor, was virtually absent. Mig was produced by the allograft endothelium as well as by recipient allograft-infiltrating macrophages and neutrophils, indicating the synergistic interactions between innate and adaptive immune compartments during acute rejection. Collectively, these results indicate that Mig is a dominant recruiting factor for alloantigen-primed T cells into cardiac allografts during acute rejection. Although Mig antagonism delays acute heart allograft rejection, the results also suggest that the alloimmune response circumvents Mig antagonism through alternative mechanisms.

Early Chemokine Cascades in Murine Cardiac Grafts Regulate T Cell Recruitment and Progression of Acute Allograft Rejection

The identification of early inflammatory events after transplant in solid tissue organ grafts that may direct T cell recruitment and promote acute allograft rejection remain largely unknown. To better understand temporal aspects of early inflammatory events in vascularized organ grafts, we tested the intragraft expression of four different chemokines in heterotopically transplanted A/J (H-2a) and syngeneic heart grafts in C57BL/6 (H-2b) recipient mice from 1.5 to 48 h after transplant. Similar temporal expression patterns and equivalent levels of chemokine expression were observed in both syngeneic and allogeneic cardiac allografts during this time period. Expression of the neutrophil chemoattractant growth-related oncogene α (KC) was observed first and reached peak levels by 6 h after transplant and was followed by the monocyte/macrophage chemoattractant protein-1 (JE) and then macrophage inflammatory proteins 1β and 1α. Administration of rabbit KC antiserum to allograft recipients within 30 min of cardiac transplantation attenuated downstream events including intra-allograft expression of the T cell chemoattractants IFN-γ-inducible protein-10 and monokine induced by IFN-γ, cellular infiltration into the allograft, and graft rejection. Similarly, depletion of recipient neutrophils at the time of transplantation significantly extended allograft survival from day 8 to 10 in control-treated recipients up to day 21 after transplant. These results indicate the induction of highly organized cascades of neutrophil and macrophage chemoattractants in cardiac grafts and support the proposal that early inflammatory events are required for optimal recruitment of T cells into allografts during the progression of acute rejection of cardiac allografts.

Neutrophils Mediate Parenchymal Tissue Necrosis and Accelerate the Rejection of Complete Major Histocompatibility Complex-Disparate Cardiac Allografts in the Absence of Interferon-γ

A major feature of acute rejection of cardiac allografts is an intense mononuclear cell infiltration accompanied by interferon (IFN)-gamma production. In the current study we tested the role of IFN-gamma in acute rejection of allografts by comparing the histopathology of rejection in wild-type versus IFN-gamma-/- recipients of major histocompatibility complex-mismatched cardiac grafts. Wild-type recipients rejected the allografts at days 8 to 9 after transplant but rejection was accelerated 2 to 3 days in IFN-gamma-deficient recipients. During rejection in wild-type recipients, the allografts were heavily infiltrated with CD8+ T cells and other mononuclear cells. In contrast, allografts in IFN-gamma-deficient recipients had few T cells but an intense neutrophil infiltration accompanied by extensive graft parenchymal necrosis. No difference in expression levels of neutrophil chemoattractants including Groalpha/KC, MIP-2, GCP-2, and MIP-1alpha, was observed in allografts retrieved from wild-type and IFN-gamma-/- recipients. Depletion of neutrophils from IFN-gamma-deficient recipients delayed rejection until days 8 to 10 after transplant and restored the histopathology of acute allograft rejection to that observed in allografts rejected by wild-type recipients. These results indicate the potent regulatory properties of IFN-gamma during acute rejection directed at neutrophil infiltration into allografts and mediating graft tissue necrosis.

Early T cell response to allografts occuring prior to alloantigen priming up-regulates innate-mediated inflammation and graft necrosis

The early inflammatory response within organ allografts is initiated by ischemia/reperfusion (I/R) and promotes subsequent alloantigen-primed T cell recruitment into and rejection of the graft. Polymorphonuclear leukocyte (PMN)-mediated tissue damage is a primary component of the early inflammation in allograft rejection. We sought to compare and elucidate the mechanism of early PMN infiltration into cardiac isografts and allografts. Despite identical production of PMN attractant chemokines, PMN infiltration following reperfusion into syngeneic and allogeneic grafts was not equivalent. PMN infiltration into isografts peaked at 9 to 12 hours post-transplant and quickly resolved. In contrast, PMN infiltration into allografts continued to elevated levels, peaking at 24 hours post-reperfusion. This amplified PMN infiltration into allografts did not resolve until 72 hours post-reperfusion and was accompanied by marked parenchymal necrosis. This early innate inflammatory response was regulated by IFN-gamma-producing CD8+ T cells present in the recipient before detectable alloantigen T cell priming. Co-culture with CD62L(low) CD8+ T cells, but not CD62L(high) CD8+ or CD62L(low) CD4+ T cells, harvested from naïve animals induced allogeneic endothelial cells to express IFN-gamma-dependent chemokines. These data demonstrate CD8+ T cell-mediated attack on the vascular endothelium of allografts within hours following organ reperfusion that amplifies innate immune-mediated intra-graft inflammation and necrosis.

Donor-specific antibody in chronic rejection is associated with glomerulopathy, thickening of peritubular capillary basement membrane, but not C4d deposition

Abstract:  The impact of post‐transplant donor‐specific antibody (DSA) on the development of chronic rejection has been focused recently. The aim of this study was to evaluate the significance of DSA, graft function and pathological factors of chronic rejection. Seventy‐three kidney recipients who underwent protocol biopsy were included in the study. The median follow‐up period after transplant was 40 months. The presence of anti‐HLA antibody (aHLAAb) and DSA was tested using flow beads analysis (FlowPRA®). The patients were divided into a group with DSA, a group with non‐donor‐specific aHLAAb and a group without aHLAAb. Protocol biopsy specimen were compared for transplant glomerulopathy (cg), vasculopathy (cv), C4d deposition at peritubular capillary (PTC), peritubular capillaritis (ptc score 0–3) and thickening of PTC basement membrane (ptcbm score 0–3) as recently proposed. The presence of DSA was significantly associated with the presence of cg, ptcbm. The group with non‐donor‐specific aHLAAb had ptcbm but did not have cg. The group without aHLAAb rarely showed ptcbm. The presence of DSA was associated with impaired graft function. C4d was not specific for the patients with DSA. These histopathological markers are useful in the detection of immunological chronic rejection. Early detection by screening tests will be important for treatment before irreversible change occurs.

Epididymo-orchitis caused by intravesically instillated bacillus Calmette-Guérin : Genetically proven using a multiplex polymerase chain reaction method

Abstract  The intravesical instillation of bacillus Calmette‐Guérin (BCG) is a standard therapy for superficial bladder carcinoma. Tuberculosis‐like inflammation in the genitourinary tract is a serious complication of BCG. It can occur after a long interval from the cessation of the intravesical BCG therapy. If inflammation occurs, it is necessary to test whether the BCG strain has caused it or another mycobacterium species has. However, there has never been a report that proves BCG causes the inflammation, because BCG is difficult to differentiate from other strains of Mycobacterium bovis and other members of the Mycobacterium tuberculosis complex by conventional tests, including regular polymerase chain reaction (PCR). We first present a case of epididymo‐orchitis, which developed 31 months after the cessation of BCG therapy, detected using a multiplex PCR method as having been caused by BCG. Our report illustrates the efficacy of this method to detect the responsible microbe that is thought to be transmitted from the instillated BCG strain.

Quadruple immunosuppression with basiliximab, tacrolimus, mycophenolate mofetil and prednisone is safe and effective for renal transplantation

Abstract:  Introduction:  Recent immunosuppression with tacrolimus and mycophenolate mofetil has improved the results of renal transplantation. In this study, we analyzed the effect and safety of basiliximab as an induction therapy.

Long-term histopathology of allografts in sensitized kidney recipients

Successful desensitization therapy has brought satisfying short‐term outcomes in the recipients with anti‐donor antibody. We analyzed the long‐term pathology of the allografts in the sensitized kidney recipients. Eleven stable recipients after desensitization against positive flow cytometry T‐cell crossmatch (FTXM) were included. They were divided into two groups, based on the protocol biopsies findings at three to eight yr (group 1: subclinical glomerulitis and/or peritubular capillaritis, n = 5 and group 2: no rejection, n = 6). Estimated glomerular filtration rate (eGFR), presence of donor‐specific antibody (DSA), mean channel shift (MCS) of FTXM, urine protein levels, acute antibody‐mediated rejection (AAMR) episodes, and protocol biopsy findings were compared. Chronic transplant glomerulopathy was found in final biopsy of all group 1 cases. DSA was positive in 60% but C4d was positive in 20% case of the group 1. The history of AAMR was only found in the group 1. There was no difference in eGFR decline or proteinuria. The MCS of FTXM was higher in the group 1. The recipients with AAMR history, high MCS in FTXM, and subclinical microvascular inflammation in the early protocol biopsies have risk for developing chronic rejection in long term.

De novo proteinuria with pathological evidence of glomerulonephritis after everolimus induction.

A 68‐year‐old man who underwent living‐unrelated kidney transplantation from his spousal donor was immunosuppressed with tacrolimus and mycophenolate mofetil. Despite his uneventful clinical course, protocol biopsy at 2 years post transplant showed de novo CNI tubulotoxicity despite low tacrolimus exposure. Everolimus was added in order to discontinue TACER. However, prominent proteinuria impeded continuation of everolimus since biopsy showed diffuse glomerular endocapillary proliferation without C4d deposition. No donor‐specific antibody was detected. Pulse steroids were given and proteinuria returned to normal with histological reversal.