Cytoplasm Of Mesangial Cells Research Articles

Inhibition of the K+ Channel KCa3.1 Reduces TGF-β1-Induced Premature Senescence, Myofibroblast Phenotype Transition and Proliferation of Mesangial Cells

ObjectiveKCa3.1 channel participates in many important cellular functions. This study planned to investigate the potential involvement of KCa3.1 channel in premature senescence, myofibroblast phenotype transition and proliferation of mesangial cells.Methods & MaterialsRat mesangial cells were cultured together with TGF-β1 (2 ng/ml) and TGF-β1 (2 ng/ml) + TRAM-34 (16 nM) separately for specified times from 0 min to 60 min. The cells without treatment served as controls. The location of KCa3.1 channels in mesangial cells was determined with Confocal laser microscope, the cell cycle of mesangial cells was assessed with flow cytometry, the protein and mRNA expression of KCa3.1, α-smooth muscle actin (α-SMA) and fibroblast-specific protein-1 (FSP-1) were detected with Western blot and RT-PCR. One-way analysis of variance (ANOVA) and Student-Newman-Keuls-q test (SNK-q) were used to do statistical analysis. Statistical significance was considered at P<0.05.ResultsKca3.1 channels were located in the cell membranes and/or in the cytoplasm of mesangial cells. The percentage of cells in G0-G1 phase and the expression of Kca3.1, α-SMA and FSP-1 were elevated under the induction of TGF-β1 when compared to the control and decreased under the induction of TGF-β1+TRAM-34 when compared to the TGF-β1 induced (P<0.05 or P<0.01).ConclusionTargeted disruption of KCa3.1 inhibits TGF-β1-induced premature aging, myofibroblast-like phenotype transdifferentiation and proliferation of mesangial cells.

Glomerular Lipidosis Accompanied by Renal Tubular Oxalosis in Wild and Laboratory-Reared Japanese Rock Ptarmigans (Lagopus mutus japonicus)

Glomerular lipidosis is a disease characterized by lipid accumulation in mesangial cells but that has not been fully investigated in avian species. We examined four wild and two laboratory-reared Japanese rock ptarmigans (Lagopus mutus japonicus)--an endangered avian species--presenting vacuolar deposits in the glomeruli. All cases had vacuolar deposits in the glomeruli. In the wild cases, fewer than 30% of all glomeruli were affected, compared with more than 90% in the laboratory-reared cases. In the wild cases, most deposits were mild and restricted to the mesangial areas of glomeruli. In the laboratory-reared cases, nearly all of the deposits covered entire glomeruli. Electron microscopy of mild deposits revealed vacuoles in the cytoplasm of mesangial cells. These vacuoles were positive for Sudan III, Sudan black B, oil red O, Nile blue, periodic acid-Schiff, Schultz test, and digitonin stain and were negative for performaric acid-Schiff stains. Based on these results, we diagnosed the glomerular lesion as glomerular lipidosis caused by uptake of low-density lipoprotein in mesangial cells. Except for one wild case, all cases exhibited renal tubular oxalosis. The severity of tubular oxalosis tended to be related to the severity of glomerular lipidosis: In cases of mild glomerular lipidosis, tubular oxalosis was also mild or absent. We therefore diagnosed the primary lesion as glomerular lipidosis accompanied by tubular oxalosis. The four wild cases came from different zones and therefore had no opportunities to interbreed and no common relatives. We believe these data support the hypothesis that glomerular lipidosis is a disease of the general population ofJapanese rock ptarmigans. This is the first report of glomerular lipidosis accompanied by renal tubular oxalosis in an avian species.

Renal expression of two 'thyroid-specific' genes: thyrotropin receptor and thyroglobulin.

Numerous renal abnormalities accompany thyroid disease, most of which have been ascribed to the effects of thyroid hormone on renal metabolism. In the present report, we investigate the renal expression of the nominally thyroid-specific proteins, thyroid-stimulating hormone (TSH) receptor (TSHR) and thyroglobulin (Tg), as potential links between renal and thyroid function. The expression of TSHR has been identified in several extrathyroidal tissues, but its presence in the kidney remains controversial. We have used reverse-transcriptase polymerase chain reaction and DNA sequencing to demonstrate the presence of TSHR transcript in human and mouse kidney, in a primary culture of human kidney, and in a green monkey kidney epithelioid cell line. Furthermore, human kidney cells responded to TSH with a 2.5- fold increase in intracellular cyclic adenosine monophosphate, suggesting the presence of functional TSHR protein. Comparison of renal expression of TSHR in a bovine growth hormone transgenic mouse model of progressive glomerulosclerosis with control mice suggested increased TSHR transcript in the renal cortex of transgenic animals. TSHR transcript was also detected in mouse mesangial cells in vitro which responded to TSH with significant increases in the formation of three-dimensional hillhocks. Polymerase chain reaction also confirmed the presence of Tg transcript in human and mouse kidneys and in mouse mesangial cells, but no effect of either TSH or cyclic adenosine monophosphate on Tg transcript levels could be discerned. Immunofluorescent staining with a monoclonal anti-Tg antibody identified positive staining in the cytoplasm of mesangial cells. These data suggest that the kidney is capable of expressing the thyroid-specific genes, TSHR and Tg, which could conceivably mediate effects of thyroid disease in the kidney.

Localization by Immunoelectron Microscopy of Schistosoma mansoni Antigens in the Glomerulus of the Hamster (Mesocricetus auratus) Kidney

Adult worm antigen (AWA) and soluble egg antigen (SEA) were localized ultrastructurally by immunoelectron microscopy using two monoclonal antibodies in the glomeruli of hamsters infected with Schistosoma mansoni cercariae or injected with S. mansoni eggs. AWA was detected in all cercaria-infected groups from the 30th day on and was present mainly in cytoplasm of mesangial cells, mesangial matrix, and glomerular basement membrane, either as isolated gold particles or in small electron-dense deposits of probable immune origin. AWA was encountered also on the inner side of the glomerular basal membrane, close to endothelial cells, and in the foot processes of the glomerular epithelial cells. SEA was detected at similar sites, apparently in lesser amounts, in uninfected hamsters inoculated with S. mansoni eggs into the jugular vein. Schistosomal antigens are apparently processed mainly by mesangial cells which are considered to be critical in the pathogenesis of S. mansoni associated glomerulopathy. Mesangioproliferative glomerulonephritis, immunoglobulin (IgG and IgM), and C₃ deposits were observed in hamsters in which AWA and SEA were visualized. During early phases of the infection and in hamsters in which granulomatous pneumonitis was induced by S. mansoni eggs, glomeruli were unchanged or showed a slight mesangial proliferation. Our findings suggests that egg antigens also contribute to the pathogenesis of experimental glomerulopathy in the hamster.

Precise ultrastructural localization of endothelial leukocyte adhesion molecule-1, vascular cell adhesion molecule-1, and intercellular adhesion molecule-1 in patients with IgA nephropathy.

Using light and electron microscopy, we performed an immunohistochemical study of endothelial leukocyte adhesion molecule-1 (ELAM-1), vascular cell adhesion molecule-1 (VCAM-1), and intercellular adhesion molecule-1 (ICAM-1) in 15 patients with IgA nephropathy to clarify the localization of these adhesion molecules. The normal portions of 2 kidneys removed due to localized carcinoma and 3 biopsies from patients without glomerular disease were used as a control. By light microscopy, ELAM-1, VCAM-1, and ICAM-1 all showed positive staining in IgA nephropathy, with the intensity of staining following the sequence ICAM-1 > VCAM-1 > ELAM-1. ELAM-1 and VCAM-1 showed a patchy distribution of moderate staining in the tissues, including the mesangium, crescents, adhesions, and tubules. In contrast, there was marked linear ICAM-1 staining throughout the vascular walls. ELAM-1 and VCAM-1 were positive on the basolateral surfaces of a few proximal tubular epithelial cells in association with inflammatory cell infiltration, while ICAM-1 was found on the brush border. ICAM-1 was positive in the glomerular capillary walls and interstitial vessels of the control kidney tissue, while ELAM-1 and VCAM-1 were virtually absent. By electron microscopy, ELAM-1 positivity on the urinary surface of the parietal/visceral epithelial cells was often associated with adherent mononuclear cells in the urinary space. VCAM-1 positivity was increased in the perinuclear space and/or cytoplasm of mesangial cells as well as at the mesangial cell-endothelial cell interface. These findings suggest that ELAM-1 and VCAM-1 may be more closely related than ICAM-1 to the major histopathological changes occurring in IgA nephropathy, including mesangial expansion, formation of crescents and adhesions, and tubulointerstitial injury.

Age-related histochemical and ultrastructural changes in renal glomerular mesangium of APA hamsters.

Syrian hamsters of the APA strain (APA hamsters) develop spontaneous mesangial thickening in the renal glomeruli from an early age. They also develop focal and segmental glomerulosclerosis (FSG) at and after 6 months of age. In this study, histopathological, histochemical and electron microscopical examinations were conducted to clarify the age-related renal glomerular changes in spontaneous FSG of APA hamsters. Histopathological analysis revealed that expansion of the mesangial region and thickening of the glomerular basement membrane (GBM) in the glomeruli of APA hamsters progressed with age. These age-related changes appeared earlier in male animals. Immunohistochemical analysis suggested that type IV collagen was responsible for the expansion of the mesangial region and thickening of the GBM. In lectin histochemical analysis, positive sites for WGA with and without neuraminidase pretreatment and PNA with neuraminidase pretreatment were detected mainly in podocytes and were expanded with age. Ultrastructurally, the increase in basement membrane-like materials in the mesangial matrix, development of intracellular organella of mesangial cells and migration of a part of the mesangial cell cytoplasm into the GBM were observed at and after 6 months of age. These features suggested that mesangial cells played an important role in the age-related expansion of the mesangial region and thickening of the GBM.

12 Renal pathology in pre-eclampsia

Pre-eclampsia affects the kidney both functionally and morphologically. Renal haemodynamics decrease and urinary protein excretion increases, in part due to lesions affecting the glomerulus, where a combination of changes produces a characteristic appearance and permits differentiation of pre-eclamptic nephropathy from other glomerular alterations associated with hypertension in pregnancy. In pre-eclampsia the glomerulus is diffusely enlarged and bloodless, due not to proliferation, but to hypertrophy of the intracapillary cells. These alterations, best described ultrastructurally, include hypertrophy of the cytoplasmic organelles in endothelial and occasionally mesangial cells, particularly the lysosomes, which undergo marked enlargement and vacuolization (due to accumulation of free neutral lipids). These reactive changes have been termed 'glomerular capillary endotheliosis'. Other lesions, observed occasionally, include subendothelial and mesangial electron-dense deposits, as well as interposition of mesangial cell cytoplasm or mesangial matrix along an otherwise normal basement membrane. Some investigators have described immunohistological findings (presence of IgM, IgG and fibrin) which they believe specific for pre-eclampsia, and others have claimed the disease may cause focal segmental glomerulosclerosis (FSGS). We believe the immunohistological findings are non-specific and insudative, and that FSGS when present predates the pre-eclamptic complication. Finally, the renal lesions appear fully reversible and the disease has no remote cardiorenal effects on its patients.

Tenascin is synthesized and secreted by rat mesangial cells in culture and is present in extracellular matrix in human glomerular diseases.

Tenascin (TN) is a large oligomeric protein recently described as a component of the extracellular matrix. The distribution of TN in adult kidney tissue has not been adequately evaluated, but preliminary data have suggested that TN is variably seen in rare mesangial areas and in stroma surrounding some tubules. The enlargement of the mesangial matrix (mesangial sclerosis) is a common feature of many renal diseases and is thought to be partially related to oversynthesis of the normal components of the mesangial matrix (collagen type IV, laminin, fibronectin, and heparan sulfate proteoglycans) by mesangial cells. However, the possibility that mesangial cells are also the source of other extracellular matrix proteins that participate in the process of mesangial sclerosis has not been explored. In this study, the synthesis of TN by cultured rat mesangial cells was documented by the following observations: (1) Northern hybridization of total RNA extracted from mesangial cells showed two distinct species of TN mRNA; (2) immunoblotting of the protein extracted from the conditioned medium demonstrated four TN protein bands; (3) immunoblotting of the protein extracted from the mesangial cell lysate demonstrated at least four TN protein bands; and (4) immunohistochemical techniques identified TN within the cytoplasm of mesangial cells and in the surrounding extracellular matrix.(ABSTRACT TRUNCATED AT 250 WORDS)

12 Renal pathology in pre-eclampsia

Pre-eclampsia affects the kidney both functionally and morphologically. Renal haemodynamics decrease and urinary protein excretion increases, in part due to lesions affecting the glomerulus, where a combination of changes produces a characteristic appearance and permits differentiation of pre-eclamptic nephropathy from other glomerular alterations associated with hypertension in pregnancy. In pre-eclampsia the glomerulus is diffusely enlarged and bloodless, due not to proliferation, but to hypertrophy of the intracapillary cells. These alterations, best described ultrastructurally, include hypertrophy of the cytoplasmic organelles in endothelial and occasionally mesangial cells, particularly the lysosomes, which undergo marked enlargement and vacuolization (due to accumulation of free neutral lipids). These reactive changes have been termed 'glomerular capillary endotheliosis'. Other lesions, observed occasionally, include subendothelial and mesangial electron-dense deposits, as well as interposition of mesangial cell cytoplasm or mesangial matrix along an otherwise normal basement membrane. Some investigators have described immunohistologic findings (presence of IgM, IgG and fibrin) which they believe specific for pre-eclampsia, and others have claimed the disease may cause focal segmental glomerulosclerosis (FSGS). We believe the immunohistologic findings are non-specific and insudative, and that FSGS when present predates the pre-eclamptic complication. Finally, the renal lesions appear fully reversible and the disease has no remote cardiorenal effects on its patients.

Glomerular changes in renal allografts.

The renal allograft is host to a number of injuries and all its structural components are prone to damage. The glomeruli respond to these varied stimuli in many ways. The fibrinoid necrosis, thrombosis, and polymorphonuclear cell exudation that accompany hyperacute or accelerated rejection are well-recognized. The transplant may also be afflicted by forms of de novo or recurrent glomerulonephritis. Apart from these, there are other patterns of reaction. The mesangium is often the site of a rapidly reversible change; it expands readily. Arterial changes initiate ischemia and collapse of glomerular capillary spaces. Glomerulitis accompanies cases of acute rejection, but when seen as a predominant feature, usually antedates chronic rejection. Heavy proteinuria may be associated with profound alterations in the peripheral capillary basal lamina including irregular thickening, interposition of mesangial cell cytoplasm, and lamellation. Allografts with these glomerular changes eventually fail.

Detection of cross-reactive anti-DNA antibody idiotypes on renal tissue-bound immunoglobulins from lupus patients.

Cross-reactive anti-DNA antibody idiotypes have been identified on tissue-bound immunoglobulins in a study of renal biopsies from 26 systemic lupus erythematosus (SLE) patients. 12 (46%) biopsies were shown to have one or both the idiotypes tested for by anti-idiotypic reagents. The idiotypes were identified in the glomerular basement membrane, the mesangial cell cytoplasm, and in focal tuft proliferations. In contrast, in none of 24 immunoglobulin-positive disease control biopsies could either idiotype be demonstrated. Blocking studies in two patients indicated that the idiotypes were on anti-DNA antibodies. These findings indicate that some tissue-bound auto-antibodies are derived from related families of high-frequency germ line genes that are expressed in SLE patients. The potential role of anti-idiotypic therapy in SLE is discussed.

Electron microscopic studies on IgA nephropathy.

We carried out electron microscopic studies on renal tissues from 9 patients with IgA nephropathy. Electron dense deposits were present in the mesangial area in all cases, subendothelial deposits in 4, and subepithelial deposits in only one. In basement membrane, segmental swelling and rarefaction of basement membrane substance were observed. In some cases the degenerated basement membrane substance protruded through the dilated endothelial fenestration into capillary lumina. Focal splitting, attenuation, mouse eaten appearance, and herniation of basement membrane were seen in a high incidence. Mesangial cells possessed well developed rough endoplasmic reticulums and polysomes. In the peripheral areas of mesangial cell cytoplasm, there was accumulation of electron dense substance and this was occasionally continuously present in the mesangial matrix. There was segmental swelling of endothelial cell cytoplasm, resulting in loss of fenestration. Epithelial cells had well developed rough endoplasmic reticulums and polysomes. Segmental foot process fusion was seen, and these processes, regardless of fusion, became electron denser in the area close to the basement membrane. Multivesiculated bodies were present in all cases in the epithelial cells and in 7 cases in the endothelial cells. Spherical microparticles were present in 3 cases in the urinary space or between the basement membrane and the epithelial cells.

Renal pathology of fetuses with congenital nephrotic syndrome of the Finnish type. 2. A qualitative and quantitative electron microscopic study.

A qualitative and quantitative transmission electron microscopic study was performed on kidneys of fetuses with congenital nephrotic syndrome of the Finnish type (CNF) and controls obtained from elective pregnancy terminations. The gestational age of the fetuses ranged from 16–22 weeks. A consistent qualitative change of CNF glomeruli was the nearly total loss of epithelial cell foot processes, with smudging of epithelial cytoplasm as broad masses on the glomerular basement membrane. Adjacent to the basement membrane there were increased amounts of fibrillar material in the epithelial cytoplasm. The number of pinocytic droplets was increased both in epithelial and endothelial cytoplasm and some of the capillary loops showed swelling of endothelial cells with poor fenestrations in cases of CNF. Mesangial hyperplasia was focally evident in the most mature CNF glomeruli with an increase of the mesangial matrix. A striking qualitative feature in CNF, only rarely seen in controls, was the occurrence of electron lucent vacuoles and heteromorphic residual bodies in the mesangial cell cytoplasm. The total thickness of the glomerular basement membrane was comparable in CNF and controls but the width of the lamina densa was significantly smaller in CNF than controls. Lamina rara interna of the basement membrane showed irregular widening with accumulation of coarse granular material. Reabsorption droplets of proximal tubular cytoplasm were significantly increased in CNF. With the dilatation of the tubule the droplets almost totally disappeared. Based on the results of the study it is suggested that the basic pathogenetic defect in CNF lies in faulty development of the lamina densa of the glomerular basement membrane and, secondary to this, various glomerular and tubular changes occur. In addition to evidence of extensive foot process fusion, mesangial cell cytoplasmic vacuoles represent a diagnostically important qualitative change in the glomeruli of fetal CNF.

Mesangial cell cytoplasm and glomerular disease

In this study we have examined mesangial cell cytoplasm as seen by light microscopy of plastic embedded renal biopsy material. An assessment of mesangial cell morphology has been made in each of the 350 renal biopsies, and this has been correlated with the disease process. A surprisingly close correlation with the type of disease present was demonstrated. Thus it appears that an increase in mesangial cell cytoplasm correlates with an active process involving the mesangial area. It is suggested that the index of mesangial cell reactivity may be useful in diagnosis of glomerular disease processes.

Recurrence of mesangial deposition of IgA after renal transplantation

Mesangial deposition of IgA is found in about 20% of adult patients with idiopathic glomerular disease in France. The deposition involves the mesangium of every glomerulus and persists during the whole course of the nephropathy. We therefore believe it to be the hallmark of the disease in these patients [1, 2]. Mesangial IgA does not contain secretory piece, and it is usually accompanied by some IgG, IgM has been found by some observers [3, 4], but not by ourselves. C3 is present without Clq or C4 [4, 5], suggesting activation of the complement system by the alternate pathway [6], This is further supported by the demonstration of properdin [4,7]. In our experience, however, the deposition of properdin is much less conspicuous than in acute glomerulonephritis or membranoproliferative glomerulonephritis. By light microscopy, the kidney usually has the appearance of focal proliferative glomerulonephritis but, with thin sections and good trichrome stains, it is often possible to see some fibrinoid deposits in the mesangium of even those glomeruli which show no focal changes. Mild diffuse mesangial hypercellularity is present in some cases, and sometimes the focal changes may be similar to those observed in focal glomerulosclerosis. In advanced cases, interstitial and arteriolar changes become prominent, and without the help of immunofluorescence microscopy, these cases may be misdiagnosed as interstitial nephritis or nephrosclerosis even by experienced pathologists. With the electron microscope, electron-dense deposits are seen in the mesangium of every glomerulus. Silver impregnation may be necessary to clearly distinguish the deposits from the basement membrane and the mesangial matrix. There may sometimes be a slight extension of the deposits to the subendothelial areas. The cytoplasm of mesangial cells is usually richer in organelles than usual but no evidence of phagocytosis of the deposits by these cells can be demonstrated. In adult patients, the disease is usually manifested by light proteinuria and microscopic hematuria. In one-half of the cases, macroscopic hematuria occurs either as a single episode or as recurrent attacks. Gross hematuria is usually precipitated by an upper respiratory tract infection or strenuous exercise. It may be accompanied by loin pain and dysuria. In some patients proteinuria is heavier, and occasionally the nephrotic syndrome may develop. Hematuria and proteinuria may disappear for variable periods of time, but renal biopsy specimens obtained during periods of clinical remission have shown that the mesangial deposits persist. The course of the disease in most adult patients is long-standing but its clinical expression is mild, with little evidence of progression over decades. Nevertheless, hypertension and renal insufficiency may occur. Terminal renal failure eventually develops in about one case in five, and unfortunately we know of no way to predict which patient will follow this course. There is no relationship between the amount of mesangial deposition, which usually remains fairly constant in a given patient, and the severity of the disease. In the child, the mesangial deposition of IgA accounts for about 10% of cases of glomerular disease. It nearly always presents as recurrent hematuria [8,9]; renal tissue often looks normal by light microscopy. The prognosis of recurrent hematuria in children is generally considered to be good, and it is therefore of note that the disease began during childhood in several patients of this report. In most patients, with mesangial deposition of IgA, the serum concentration of IgA is high, but in some patients, it may be normal or even low. The serum concentrations of IgG, IgM and C3 are normal or slightly elevated. There are still no clues as to the etiology of this disease.