Detection of urinary macrophages expressing the CD16 (Fc gamma RIII) molecule: a novel marker of acute inflammatory glomerular injury

BACKGROUND: The CD16 antigen is the Fc gamma receptor III. CD14+CD16+ cells are proinflammatory monocytes/macrophages (Mo/M phi) that constitute a minor population in the peripheral blood of healthy individuals. Little is known about the expression of CD16 antigen on Mo/M phi in glomerulonephritis. METHODS: Flow cytometric analyses were performed on urine and blood samples obtained from 209 patients with various renal diseases. Patients variously suffered from rapidly progressive crescentic glomerulonephritis (RPGN), membranoproliferative glomerulonephritis (MPGN), postinfectious acute glomerulonephritis (AGN), Henoch-Sch?nlein purpura nephritis (HSPN), IgA nephropathy (IgAN), membranous nephropathy (MN), minimal change nephrotic syndrome (MCNS), lupus nephritis (LN), acute interstitial nephritis, hereditary nephropathy, idiopathic renal hematuria (IRH), and renal stone. RESULTS: The CD16+ M phi population of cells was present in the urine of hematuria-positive patients with proliferative glomerulonephritis, including AGN, IgAN, RPGN, MPGN, and LN with acute inflammatory lesions, such as endocapillary proliferation, tuft necrosis, and cellular crescents. In contrast, the urinary CD16+ M phi population was negligible in hematuria-positive patients with nonproliferative renal disease, including hereditary nephropathy, IRH, and renal stone and also in patients with proliferative glomerulonephritis lacking acute inflammatory lesions. Total urinary M phi of these patients were much less than those of patients having proliferative glomerulonephritis with acute inflammatory lesions. Transient expansion of the CD16+ M phi population in urine was observed during the acute exacerbation of urinary abnormalities, whereas the disappearance of CD16+ M phi closely preceded the amelioration of urinary abnormalities in patients with proliferative glomerulonephritis. In 38 of the 98 patients positive for CD16+ M phi population in urine, the CD16+ Mo population was negligible in peripheral blood. Immunohistochemically, CD16+ M phi were present in the glomeruli of active proliferative glomerulonephritis, whereas such cells were absent in inactive proliferative glomerulonephritis or nonproliferative glomerular diseases. CONCLUSION: CD16+ M phi may be effector cells involved in the acute inflammation common to all types of proliferative glomerulonephritis. Furthermore, the detection of CD16+ M phi in urine, as well as urinary M phi counts, may serve as a useful indicator of the active stage of proliferative glomerulonephritis.     

Monoclonal antibody analysis of glomerular, tubulo-interstitial infiltrating immune cells in various glomerulonephritis

To investigate the role of cell-mediated immunity (CMI) in glomerulonephritis (GN), we identified the infiltrating immune cells both within the glomerulus and in the interstitium. Frozen sections from 103 patients with various forms of GN: 10 with minor glomerular abnormality (MGA) as control, 10 with minimal change nephrotic syndrome (MCNS), 10 with membranous nephropathy (MN), 9 with focal glomerulosclerosis (FGS), 30 with IgA nephropathy (IgAN), 22 with acute post streptococcal glomerulonephritis (APSGN), and 2 with rapidly progressive glomerulonephritis (RPGN) were examined using monoclonal antibodies (MoAb) by indirect immunoalkaline-phosphatase labelling. In most glomerulonephritis, monocyte/M phi and helper/inducer T cells were predominantly infiltrating in the interstitium, but intraglomerular infiltration was rare, except for APSGN. This interstitial infiltration increased proportionally to the level of serum creatinine, and was most prominent in RPGN. Apparently different distribution was seen in APSGN, that is, prominent increase in total number of intra-glomerular monocyte/M phi infiltration with slightly increased T cells. The change was correlated with time after onset; namely the more leucocytic infiltration was observed when the tissue was taken earlier. These data suggest that in APSGN, monocyte/M phi accumulate in glomeruli via cell mediated immunity in addition to humoral immune mechanism resulting in glomerular hypercellularity, whereas in most chronic glomerulonephritis interstitial leucocyte infiltration, particularly helper T cells and monocyte/M phi may play an important role in the progression of glomerulonephritis.     

胃癌组织中HIF-1α,CD68及CD206在胃癌及癌旁组织中的表达及临床意义

目的 分析低氧诱导分子-1α（HIF-1α）及肿瘤相关巨噬细胞（TAM）相关抗原CD68、CD206在胃癌及癌旁组织中的表达情况。方法 利用免疫组化技术检测43例胃癌和癌旁组织中HIF-1α、CD68、CD206的表达状态,计算三种蛋白表达的阳性率及阳性细胞数,揭示其与临床因素的相关性。结果 HIF-1α、CD68、CD206的阳性表达率分别为58.1%、69.8%、51.2%,均高于癌旁组织（P<0.05）。胃癌、癌旁组织中每个视野下CD68⁺细胞数分别为（39.7±7.6）、（8.5±2.8）个;CD206⁺阳性细胞数分别为（32.0±9.2）、（3.4±1.8）个;HIF-1α⁺细胞数（22.9±5.6）、（2.1±1.2）个;组间比较差异均有统计学意义（P<0.01）。胃癌组织中CD206⁺细胞数与HIF-1α⁺细胞数表达呈正相关（P<0.01,R²=0.641）。三种蛋白的表达与胃癌病理分期、淋巴结转移明显相关（P<0.05）。结论 胃癌组织中CD68、CD206、HIF-1α表达率及阳性细胞数明显增加,且CD206与HIF-1α表达呈正相关。胃癌缺氧区域对M2型巨噬细胞有趋化作用,促进胃癌发生发展。     

Clinical significance of costimulatory molecules CD80/CD86 expression in IgA nephropathy

BACKGROUND: IgA nephropathy (IgAN) is the most common form of human glomerulonephritis. Tubulointerstitial inflammation with infiltration of mononuclear cells plays an important role in the progression of IgAN. Activation of T cells requires costimulatory signals through binding of CD28 receptor with cognate ligands (CD80/CD86) located on antigen-presenting cells (APC). To assess the clinical significance of this regulatory pathway participation in the pathogenesis of IgAN, a comprehensive immunohistologic evaluation was conducted on renal tissue of IgAN in different phases of progressive injury. METHODS: Thirty-three cases of IgAN and ten cases of non-IgA mesangial proliferative glomerulonephritis (PGN) with minor tissue damage as controls were investigated. Monoclonal antibodies were used to assess the expression of CD80, CD86, CD68, CD14, CD45RO, human leukocyte antigen-DR (HLA-DR), and intercellular adhesion molecule-1 (ICAM-1) in renal tissues. Clinical and expression data were compared at the time of renal biopsy. RESULTS: CD80+ and CD86+ cells were observed more in IgAN patients with progressive renal injury than in mild cases and controls. CD80 was limited to tubular epithelial cells and was complemented by HLA-DR expression. CD86 was expressed in the glomerulus, periglomerular area, and peritubular interstitium. Activated T cells (CD45RO+), monocytes (CD14+), macrophages (CD68+), and CD86 showed similar distributions. Positive correlations were found between CD86+ cells and CD45RO, CD14, and CD68 positive cells and between CD80+ tubuli and peritubular interstitial CD45RO+ cells. The number of interstitial CD86 positive cells and the percentage of CD80+ tubuli were correlated with renal function. Most CD86+ cells were monocyte/macrophages. CONCLUSION: This study suggested that CD80 and CD86 activate T cells in IgAN, CD80/CD86 expressions correlated with renal function at the time of renal biopsy, and monocyte/macrophages and tubular epithelial cells act as APC.     

Renal expression of matrix metalloproteinases in human ANCA-associated glomerulonephritis.

BACKGROUND: Expression of matrix metalloproteinases (MMPs) by infiltrating and intrinsic renal cells is increased in inflammatory conditions, and may correlate with disease activity of glomerulonephritis. We analysed renal expression of MMPs, tissue inhibitor of metalloproteinase-1 (TIMP-1) and markers of neutrophil and monocyte infiltration in renal biopsies of patients with active anti-neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis. METHODS: Immunohistochemical expression of MMP-2, -3, -9, TIMP-1, the neutrophil- and monocyte-derived MMP activators cathepsin G, neutrophil elastase and myeloperoxidase (MPO), and the monocyte marker CD14 was determined in renal biopsies of active proteinase 3 (PR3)-ANCA (n = 7) and MPO-ANCA (n = 6) associated glomerulonephritis, and in normal renal tissue (n = 4). Double labelling experiments of MMPs and TIMP-1 were performed with MPO and CD68, labelling neutrophils and macrophages. RESULTS: MMP-2-, MMP-3-, MMP-9- and TIMP-1-positive cells were detected in ANCA-associated glomerulonephritis in glomeruli with active inflammation (cellular crescents or fibrinoid necrosis), only occasionally in normal appearing glomeruli, and not in sclerotic glomeruli and positive cells were found in the tubulo-interstitium. MMPs and TIMP-1 were expressed predominantly by MPO-and CD68-positive cells. In normal renal tissue, no expression was detected, with the exception of weak mesangial staining for MMP-2. In ANCA-associated glomerulonephritis, glomerular MMP-2, -9 and TIMP-1 correlated with glomerular cathepsin G expression, while the number of MMP-9-expressing cells per glomerulus correlated with the percentage of crescentic glomeruli. Tubulo-interstitial expression of MMPs correlated with all markers of neutrophil and monocyte infiltration, and interstitial MMP-9 and TIMP-1 expression correlated with renal function at the time of renal biopsy. CONCLUSIONS: Expression of glomerular and interstitial MMP-2, -3, -9 and TIMP-1 is increased in active ANCA-associated glomerulonephritis and correlates with inflammatory activity.     

Blockade of Macrophage Colony-Stimulating Factor Reduces Macrophage Proliferation and Accumulation in Renal Allograft Rejection

Macrophage accumulation within an acutely rejecting allograft occurs by recruitment and local proliferation. To determine the importance of M-CSF in driving macrophage proliferation during acute rejection, we blocked the M-CSF receptor, c-fms, in a mouse model of acute renal allograft rejection. C57BL/6 mouse kidneys (allografts, n = 20) or BALB/c kidneys (isografts, n = 5) were transplanted into BALB/c mice. Anti-c-fms antibody (AFS98) or control Ig (50 mg/kg/day, i.p.) was given daily to allografts from days 0-5. All mice were killed day 6 postoperatively. Expression of the M-CSF receptor, c-fms, was restricted to infiltrating CD68+ macrophages. Blockade of c-fms reduced proliferating (CD68+/BrdU+) macrophages by 82% (1.1 v 6.2%, p < 0.001), interstitial CD68+ macrophage accumulation by 53% (595 v 1270/mm2, p < 0.001), and glomerular CD68+ macrophage accumulation by 71% (0.73 V 2.48 CD68+ cells per glomerulus, p < 0.001). Parameters of T-cell involvement (intragraft CD4+, CD8+ and CD25+ lymphocyte numbers) were not affected. The severity of tubulointerstitial rejection was reduced in the treatment group as shown by decreased tubulitis and tubular cell proliferation. Macrophage proliferation during acute allograft rejection is dependent on the interaction of M-CSF with its receptor c-fms. This pathway plays a significant and specific role in the accumulation of macrophages within a rejecting renal allograft.     

CD+68细胞在原发性肝细胞癌及肝硬化组织中的分布及其意义

目的：研究CD^＋68细胞在原发性肝细胞癌（HCC）、癌旁、肝硬化及正常肝组织中的数量变化及其临床意义。方法：HCC60例，单纯性肝硬化62例，正常肝组织23例，以免疫组化SP法染色。对阳性细胞数进行定量分析。结果：（1）CD^＋68细胞平均数从高到低为：癌旁组织，肝硬化，正常肝，癌组织（P〈0．01）；（2）癌组织中CD^＋68细胞随HCC分化程度降低而减少（P〈0.05）；（3）癌组织中和癌旁组织中CD^＋68分布与临床TNM分期无关；（4）肝癌组织中15月内有转移组的CD^＋68细胞数少于无转移组（P〈0．05）。结论：CD^＋68细胞可成为反映机体抗肿瘤免疫状态及判断患者预后的重要指标之一。     

Important role for macrophages in induction of crescentic anti-GBM glomerulonephritis in WKY rats.

BACKGROUND: A crucial role for CD8(+) cells in induction of crescentic anti-glomerular basement membrane (GBM) glomerulonephritis (GN) in WKY rats was demonstrated in studies showing that depletion of CD8(+) cells completely suppressed glomerular accumulation of monocytes/macrophages (Mo/Mphi), crescent formation and proteinuria. Because these studies did not definitively identify CD8(+) cells as the cause of tissue injury, we examined the roles of Mo/Mphi in the development of anti-GBM GN. METHODS: We examined correlations between the amount of urinary protein and the numbers of glomerular CD8(+) cells or Mo/Mphi in rats after administrating different doses of anti-GBM antibody (5.0, 7.5, 10.0 and 25.0 microl/100 g body weight). The roles of Mo/Mphi in induction of GN were examined in animals by depleting Mo/Mphi in the glomerulus. To do this, rats were injected intravenously with liposome-encapsulated dichloromethylene diphosphonate (liposome-MDP) from day 3 to day 7 after anti-GBM antibody injection and they were then sacrificed at day 8. RESULTS: Liposome-MDP treatment significantly reduced the number of ED-1(+) Mo/Mphi accumulated in glomeruli from 32.1 +/- 1.2 to 1.4 +/- 0.3/glomerular cross-section (mean +/- SD, P < 0.01), and the amount of urinary protein from 103.8 +/- 19.8 to 31.8 +/- 15.9 mg/day (P < 0.01), as well as the incidence of crescentic glomeruli from 91.3 +/- 2.7 to 23.3 +/- 7.6% (P < 0.01) at day 8. This treatment also reduced the number of CD8(+) cells accumulating in the glomeruli from 5.4 +/- 0.7 to 0.5 +/- 0.1/glomerular cross-section (P < 0.01). Upregulation of glomerular intercellular adhesion molecule 1 (ICAM-1) and monocyte chemoattractant protein 1 (MCP-1) mRNA expression was suppressed by Mo/Mphi depletion. CONCLUSION: These results indicate that Mo/Mphi play an important role in the induction of crescentic anti-GBM GN and glomerular injury.     

Compartment specific expression of dendritic cell markers in human glomerulonephritis

Macrophages and dendritic cells are heterogenous and highly plastic bone marrow-derived cells that play major roles in renal diseases. We characterized these cells using immunohistochemistry in 55 renal biopsies from control patients or patients with glomerulonephritis as an initial step towards postulating specific roles for these cells in kidney disease. In proliferative glomerulonephritis numerous CD68 positive (pan monocyte, macrophage and dendritic marker) cells were found in both glomeruli and the tubulointerstitial space, however, a myeloid dendritic cell marker (DC-SIGN) was identified only in the tubulointerstitium. A significant number of plasmacytoid dendritic cells (identified as BDCA-2 positive cells) were seen at sites of interstitial inflammation, including follicular aggregates of inflammatory cells. Langerin positive cells (a marker of Langerhans' cells) were detectable but rare. The area of either CD68 or DC-SIGN positive interstitial cells correlated with serum creatinine. Low levels of DC-SIGN, DC-LAMP and MHC class II mRNA were present in the tubulointerstitial space in controls and increased only in that region in proliferative glomerulonephritis. We demonstrate that the CD68 positive cells infiltrating the glomerulus lack dendritic cell markers (reflecting macrophages), whereas in the tubulointerstitial space the majority of CD68 positive cells are also DC-SIGN positive (reflecting myeloid dendritic cells). Their number correlated with serum creatinine, which further emphasizes the significance of interstitial DCs in progressive glomerular diseases.     

Excretion of epidermal growth factor-like material in acute Henoch-Schönlein purpura nephritis

In Henoch-Schönlein purpura nephritis (HSPN), glomeruli may develop cellular crescents composed of infiltrating monocytes and proliferating renal epithelia. In this study, we demonstrate that peripheral human monocytes can release an epidermal growth factor (EGF)-like substance detectable by a radioreceptor assay, which recognizes both EGF and transforming growth factor-alpha (TGF-alpha), but not with a radioimmunoassay, which recognizes only EGF. Furthermore, we report that urine from pediatric patients during the acute phase of HSPN contains a similar EGF-like species in addition to the endogenous EGF which is normally present. The EGF-like material was not present in urine from nine healthy children or from six children with acute post-streptococcal glomerulonephritis. The extent of crescent formation in our patients is uncertain, since renal biopsy was performed in only one case. However, we speculate that the urinary material resembling TGF-alpha which appears during the acute phase of HSPN may derive from monocytes infiltrating the kidney.     

胎儿皮肤免疫细胞CD68、CD3的表达与无瘢痕愈合的关系

目的探讨人胎儿皮肤、正常成人皮肤及增生性瘢痕(HS)组织免疫细胞CD68、CD3与无瘢痕愈合的关系。方法选择笔者单位引产的16~33周胎龄的10例胎儿皮肤、7例正常成人皮肤以及18例HS标本,用免疫组织化学的方法分别检测其巨噬细胞、T淋巴细胞的表面标志CD68、CD3的表达。结果胎儿皮肤中CD68[(5±6)个/400倍视野]显著少于成人[(23±4)个/400倍视野,P<0.01],成人皮肤中CD68又显著少于HS[(38±16)个/400倍视野,P<0·01].随胎龄增加,CD68逐渐增多,在24~28周胎龄时迅速上升,28周以后上升缓慢。发育各期胎儿皮肤中均未见CD3;成人皮肤中可见少量CD3[(24±8)个/400倍视野],主要分布于表皮基底层;HS组织中CD3较多[(69±25)个/400倍视野],常聚集成片状,主要分布于真皮乳头层,在小血管周围呈袖套状分布,数量和染色强度大于成人皮肤(P<0.01).结论胎儿皮肤中CD68数量少可能与无瘢痕愈合存在一定的关系;同时胎儿皮肤无瘢痕愈合可能与胎儿皮肤中缺乏CD3有关。     

Analysis of macrophages in urine sediments in children with IgA nephropathy

AIM: Although infiltrating macrophages found in renal biopsy specimens have been accepted as a useful marker for evaluating the activity of IgA nephropathy (IgAN), it is difficult to perform renal biopsies repeatedly, especially in children. To establish a more convenient and noninvasive method for estimating the degree of macrophage infiltration we examined the number of macrophages in urinary sediments. PATIENTS AND METHODS: Ten ml of morning urine were collected from 30 children with IgAN, 10 with thin basement membrane disease (TBMD), 8 with idiopathic renal hemorrhage (IRH) which was defined as nonglomerular hematuria due to nutcracker phenomenon revealed on ultrasonography, and 10 healthy children as controls. Ten of the 30 children with IgAN were treated with combination therapy comprising prednisolone, warfarin and dipyridamole and urine samples were collected weekly during the period of treatment. Two microl of the urine sediment were smeared on glass slides, dried and stained with a monoclonal antibody to human macrophages (anti-CD68, PG-M1) followed by a FITC-conjugated secondary antibody. After staining with propidium iodide (PI), the cells were examined by fluorescence microscopy with cells stained with both FITC and PI being counted as macrophages. In addition, anti-CD68 staining was used to quantify macrophage infiltration in renal biopsies from the same group of IgAN patients. RESULTS: The number of urine macrophages in children with IgAN was significantly higher than in children with TBMD and IRH as well as the control group (p < 0.01), whereas that was similar among TBMD, IRH and healthy children. In IgAN, there was a significant correlation between urine macrophage number and the activity index (p < 0.01), proteinuria (p < 0.01) and urine WBC count (p < 0.01). In addition, there was also a significant correlation between urine macrophage number and glomerular (p < 0.05) as well as interstitial macrophage infiltration (p < 0.01). In children with IgAN who received combination therapy, urine macrophage number decreased significantly (p < 0.01) in the 1st week of treatment whilst the degree of proteinuria decreased significantly (p < 0.01) in the 4th week. CONCLUSION: Urinary macrophage number may represent a noninvasive and straightforward estimate of the pathological activity evident in renal biopsy specimens, and may also be a more sensitive indicator than proteinuria of the therapeutic effect of interventional treatments in childhood IgAN.     

Down regulation of CD45 expression on CD4 T cells during acute renal allograft rejection: Evidence of a decline in T suppressor/inducer activity

Acute rejection is associated with the activation of helper and cytotoxic cells. A shifting balance between the suppressor/inducer CD45+ CD4+ and T helper/inducer (CD4+CD45–) cells may be responsible for the transition from quiescence to overt rejection. We examined the kinetics of CD45 expression on CD4+ T cells in renal allograft recipients from pretransplant values to acute rejection and after reversal of rejection, searching for a shift in balance between helper/inducer and suppressor/inducer cell subsets. Using two color flow cytometry, the peripheral blood levels of CD4+, CD4+CD45– [T helper/inducer (Thi)], CD4+CD45+ [T suppressor/inducer (Tsi)], CD3+, and CD8+ T cells subsets and their interrelationships, were determined in 49 patients prior to transplantation, and in 10 of them, during acute rejection and after its reversal. Results were analyzed and compared to data obtained from 10 healthy blood donors. Acute rejection was associated with a significant decline in CD45+ CD4+ expression compared to quiescent phase (22% ± 3.7%vs. 26.5% ± 3.2%, p = 0.05) and controls (29.5% ± 6.2%, p = 0.01). No difference was observed compared to pretransplant levels (19.9% ± 3.2%, p = ns). CD45–/CD45+ (Thi/Tsi) ratio was lowest during quiescence (0.75) compared to rejection (0.97, p = 0.05), in controls (0.98, p = 0.05) and pretransplant values (1.4, p = 0.01). Acute rejection was characterized by higher Thi/CD8+ and lower Tsi/CD8+ ratio (103 and 88 respectively, p = 0.045), compared to clinical quiescence (104 and 116 respectively, p = 0.039). These data suggest that acute rejection is associated with down regulation of CD4+CD45+ suppressor/inducer subset. This shift may account for the transition from quiescence to overt rejection, concurring with reports on CD4+CD45 regulatory function.     

Characterization of interstitial infiltrating cells in Berger's disease

The role of infiltrating blood-borne cells in the pathogenesis of renal damage in human glomerulonephritis is under active investigation. We have evaluated leukocyte infiltrates (number of cells/mm2) in the renal interstitium of 21 patients with Berger's disease and eight normal kidneys with monoclonal antibodies and a four-layer immunoperoxidase technique. In our population of patients, the number of infiltrating T-lymphocytes (OKT11+ cells) was significantly higher (median, 132) than in the normal kidneys (median, 60). This increase was mainly due to T-suppressor/cytotoxic lymphocytes (OKT8+ cells; median, 68), while T-helper/inducer lymphocytes (Leu 3A+ cells) and monocytes were in the normal range. T-lymphocyte infiltration was more marked in ten patients with impaired glomerular filtration rate (GFR) at the time of biopsy (median, 167) than in patients with normal GFR (median, 88). In addition, ten patients who showed deterioration of renal function during the subsequent follow-up, whatever their serum creatinine levels at the time of biopsy, had significantly more total T cells (median, 269), OKT8+ cells (median, 143), and Leu 3A+ cells (median, 105) than 11 patients with persistently stable GFR and normal controls. More data are necessary to establish whether this T-lymphocyte infiltration is the consequence of a cell-mediated mechanism acting in the interstitium, concomitant with the immune-complex-mediated mechanism acting in the glomerulus, or is a nonspecific consequence of the tubulointerstitial damage induced by the immunologically mediated glomerular disease.     

Analysis of mononuclear cells in urine using flow cytometry: a useful tool for monitoring disease activity of proliferative glomerulonephritis]

We investigated CD56+ cells (natural killer cells), CD14+ cells (macrophage) and CD3+ cells (pan T cells) in urine using flow cytometry in various renal diseases including idiopathic crescentic glomerulonephritis, IgA nephropathy, membranoproliferative glomerulonephritis membranous nephropathy, pyelonephritis and idiopathic renal hematuria. A remarkable increase of CD56 cells in urine was observed in glomerulonephritis with marked necrotizing/and or crescentic (NC) lesions. The ratio of CD56+ cells/CD3+ cells was correlated with the severity of NC lesions. The ratio of CD14+ cells/CD3+ cells was increased in glomerulonephritis with endocapillary proliferation. Using immunohistochemical methods, CD56+ cells were also observed to be present in cellular crescents in biopsy specimens. These observations suggest that an analysis of these cells in urine using flow cytometry may be a useful tool for monitoring disease activity in proliferative glomerulonephritis.     

Infiltration of CD3+ and CD68+ cells in bladder cancer is subtype specific and affects the outcome of patients with muscle-invasive tumors

ObjectivesUrothelial carcinoma (UC) aggressiveness is determined by tumor inherent molecular characteristics, such as molecular subtypes, as well as by host reactions directed toward the tumor. Cell types responsible for the host?s response include tumor-infiltrating lymphocytes (TILs) and tumor-associated macrophages (TAMs). The aim of the present investigation was to explore the immunological response in relation to UC molecular subtypes and to evaluate the prognostic effect of TIL and TAM counts in tissue sections from muscle-invasive (MI) tumors.Methods and materialsTissue microarrays with 296 tumors spanning all pathological stages and grades were analyzed with antibodies for CD3, CD8, FOXP3, CD68, and CD163. Cases were classified into the following molecular subtypes: urobasal, genomically unstable, and squamous cell carcinoma–like using a combination of immunohistochemistry and histology. The Cox regression and Kaplan-Meier analyses were performed with progression-free survival and disease-specific survival as end points.ResultsUC molecular subtypes demonstrate different degrees of immunological responses; the urobasal subtype induces a weak response, the genomically unstable subtype induces an intermediate response, and the squamous cell carcinoma–like subtype induces a strong response. These subtype specific responses are independent of tumor stage and include both TILs and TAMs. The presence of infiltrating CD3⁺ TILs was significantly associated with good prognosis in the MI cases (P<0.01). This positive association was modulated by the presence of CD68⁺ TAMs. The strongest association with poor survival was observed for a high ratio between CD68 and CD3 (P = 7×10^?5).ConclusionUC molecular subtypes induce immunological responses at different levels. A high CD68/CD3 ratio identifies a bad prognosis group among MI UC cases.     

Local leukocyte proliferation as a target for cyclophosphamide in the treatment of Henoch‐Schönlein purpura nephritis grade VI

Henoch‐Schönlein purpura nephritis (HSPN) is one of the most common types of chronic glomerulonephritis in children; however, there have been few reports on the pathogenesis and management of grade VI HSPN. We present the case of a 6‐year‐old boy with grade VI HSPN accompanied by severe nephrotic syndrome and hypocomplementaemia. Immunohistological studies revealed profound glomerular accumulation of CD45‐ and CD68‐positive inflammatory cells. Moreover, some cells expressed the proliferating marker proliferating cell nuclear antigen. His proteinuria and general oedema persisted despite repeated high‐dose steroid therapy; however, these clinical symptoms immediately improved after beginning treatment with cyclophosphamide (CyP). Grade VI HSPN was successfully treated with steroids and immunosuppressants. Among immunosuppressive drugs, CyP was considered the most effective.     

The clinical relevance of plasma CD147/basigin in biopsy-proven kidney diseases

Background

Precise understanding of kidney disease activity is needed to design therapeutic strategies. CD147/basigin is involved in the pathogenesis of acute kidney injury and renal fibrosis through inflammatory cell infiltration. The present study examined the clinical relevance of CD147 in biopsy-proven kidney diseases that lead to the progression of chronic kidney disease.

Methods

Kidney biopsy specimens and plasma and urine samples were obtained from patients with kidney diseases, including IgA nephropathy (IgAN), Henoch–Schönlein purpura nephritis (HSPN), diabetic kidney disease (DKD), focal segmental glomerulosclerosis (FSGS), and membranous nephropathy (MN), who underwent renal biopsy between 2011 and 2014. Plasma and urinary CD147 levels were measured and evaluated for their ability to reflect histological features. Disease activity of IgAN tissues was evaluated according to the Oxford classification and the Japanese histological grading system.

Results

In biopsy tissues, CD147 induction was detected in injured lesions representing renal inflammation. Plasma CD147 values correlated with eGFR in patients with inflammation-related kidney diseases such as IgAN, HSPN, and DKD. Particularly in IgAN patients, plasma CD147 levels were correlated with injured regions comprising more than 50% of glomeruli or with tubular atrophy/interstitial injury in biopsy tissues. Proteinuria showed a closer correlation with urinary values of CD147 and L-FABP. Of note, plasma and urinary CD147 levels showed a strong correlation with eGFR or proteinuria, respectively, only in DKD patients.

Conclusion

Evaluation of plasma and urinary CD147 levels might provide key insights for the understanding of the activity of various kidney diseases.

     

Anti-perforin antibody treatment ameliorates experimental crescentic glomerulonephritis in WKY rats

The depletion of CD8+ cells has been shown to prevent the initiation and progression of antiglomerular basement membrane (GBM) crescentic glomerulonephritis (GN) in Wistar-Kyoto (WKY) rats. In this study, we asked whether CD8+ cells produce their effects by perforin/granzyme-mediated or by Fas ligand (FasL)-mediated pathways. The glomerular mRNA expression of perforin and granzyme B corresponded with the number of CD8+ cells, whereas that of granzyme A, Fas, and FasL did not. The enhanced mRNA level of perforin and granzyme B was not evident in CD8+-depleted rats. The number of apoptotic cells in the glomeruli was significantly increased at day 3. Perforin mRNA was found in cells infiltrating the glomerulus by in situ hybridization and by using dual-staining immunohistochemistry perforin protein was found in glomerular CD8+ cells. We found that perforin was readily visualized at the inner surface of the glomerular capillaries by immunoelectron microscopy. Based on these results, we treated animals with a perforin antibody in vivo and found that it significantly reduced the amount of proteinuria, frequency of crescentic glomeruli, and the number of glomerular monocytes and macrophages, although the number of glomerular CD8+ cells was not changed. Our results suggest that CD8+ cells play a role in glomerular injury as effector cells in part through a perforin/granzyme-mediated pathway in the anti-GBM WKY rat model of crescentic GN.