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PubMed1992

Clusterin (SGP-2) in epididymal luminal fluid and its association with epididymal spermatozoa in androgen-deprived rats.

Clusterin is a heterodimeric glycoprotein synthesized and secreted by rat Sertoli cells and epididymal epithelium. The goal of this study was to determine the presence of clusterin in the luminal fluid of the cauda epididymides and its association with the membranes of developing spermatozoa in the presence and absence of androgen. We have previously demonstrated by two-dimensional (2-D) Western blot probing for clusterin that in epididymal fluid the amounts of clusterin were: caput greater than corpus greater than cauda. Luminal fluid from cauda epididymides was collected from control and orchiectomized rats (6 and 12 days) and orchiectomized animals that received testosterone implants. Equal volumes of fluid were analyzed by 2-D Western blot probing for clusterin. Following orchiectomy, there was an increase in clusterin in the luminal fluid after 6 days and maximal amount after 12 days compared with control cauda fluid. Orchiectomized animals which received testosterone treatment showed levels of clusterin comparable to that of controls. Serum clusterin was detected in fluid of orchiectomized animals with and without testosterone. Western blots of cauda sperm membrane extracts of control animals and orchiectomized animals treated with testosterone had a very low level of epididymal clusterin, whereas extracts collected from orchiectomized animals revealed high levels of clusterin. We suggest that, in the normal animal, clusterin is secreted into the lumen of the proximal epididymis where it binds to the sperm membrane. In the distal epididymis, clusterin dissociates from sperm and is processed (proteolysis/endocytosis). We hypothesize that, in the absence of androgen, the processing and regulation of clusterin is disrupted.

Animals↗
PubMed2000

Antisense oligonucleotide of clusterin mRNA induces apoptotic cell death and prevents adhesion of rat ASC-17D Sertoli cells.

Clusterin has been known to play important roles in cell-cell and/or cell-substratum interactions. Recently we reported the transient expression of clusterin in pancreatic endocrine cells during the early developmental stages and suggested a role in aggregating the endocrine cells for islet formation. In the present study, we have investigated the involvement of clusterin in cell-substratum interaction by the inhibition of clusterin synthesis using antisense oligonucleotide. The expression of clusterin was transiently increased as early as 2-8 h after plating the ASC-17D Sertoli cells to the culture flask, which was the period of cell attachment. In addition, up-regulation of clusterin mRNA was so much greater when the Sertoli cells were plated on the petri dish for the bacterial culture instead of in a animal cell culture flask that therefore, the cells failed to attach to it. These findings suggested that interruption of cell to plate substratum interaction might lead to over-expression of clusterin from Sertoli cells to induce cell to cell aggregation or, perhaps, to re-establish attachment with the substratum. Transfection of ASC-17D Sertoli cells with a 20-base antisense oligonucleotide against clusterin mRNA resulted in extracellular release of LDH and DNA fragmentation. Sertoli cell death by antisense oligonucleotide of clusterin was sequence specific and dose dependent. Treatment of antisense oligonucleotide induced a marked reduction of synthesis for clusterin protein, but not for clusterin mRNA expression, suggesting the translational suppression of clusterin by antisense oligonucleotide. Further, microscopic observation showed that more noticeable cell death was induced by treating the antisense prior to plating the cells than by treating after cell attachment to the plate. From these results, we speculate that down-regulation of clusterin expression in the anchorage-dependent Sertoli cells prevents them from attaching to the plate, and therefore induces cell death.

Animals↗
PubMed2000

Overexpression of clusterin in human breast carcinoma.

Clusterin has been implicated in numerous processes including active cell death, immune regulation, cell adhesion and morphological transformation. The purpose of this study was to examine clusterin expression in a large series of breast carcinomas by immunohistochemistry and in situ hybridization. The study included 40 samples of non-neoplastic glandular epithelia, 42 benign lesions, 15 atypical intraductal hyperplasias, 35 carcinomas in situ, 114 invasive carcinomas, and lymph node metastases from 40 patients. Epithelial normal cells were always negative for clusterin expression and only 19% of the benign lesions presented positive staining. In contrast to the benign lesions, however, the frequency of clusterin positive samples increased in atypical hyperplasias (47%, P = 0.08), intraductal carcinomas (49%, P = 0.01) and invasive carcinomas (53%, P < 0.001). Positive staining presented a cytoplasmic pattern, except in 3 cases of invasive carcinomas which had nuclear staining. Clusterin mRNA by in situ hybridization confirmed the specific cellular pattern of clusterin expression by immunohistochemistry. Clusterin expression was associated with large tumor size (P = 0.04), estrogen and progesterone receptor negative status (P = 0.02 and P = 0.001, respectively) and with the progression from primary carcinoma to metastatic carcinoma in lymph nodes (80% metastatic nodes had positive expression) (P = 0.004). Ten of 15 (67%) primary carcinomas without clusterin expression became positive in lymph node metastases, while most (22 of 25, 88%) of the clusterin-positive primary carcinomas were also immunoreactive in metastases. In survival analysis, clusterin expression did not represent a prognostic indicator by uni- or multivariate analysis. The increased clusterin expression in breast carcinomas tended to correlate inversely with the apoptotic index (P = 0.09) which indicates that clusterin gene expression is not a prerequisite to cellular death by apoptosis. From these results, we suggest that clusterin may have a role in tumorigenesis and progression of human breast carcinomas.

Adult↗
PubMed1997

Effects of clusterin overexpression on TNFalpha- and TGFbeta-mediated death of L929 cells.

Clusterin is a widely distributed and highly conserved protein for which many functions have been proposed. We used transfected L929 cells to study the effect of clusterin expression on the regulation of cell death signals. We showed that high levels of clusterin expression, about 0.2 pg clusterin secreted per cell per 48 h period, specifically protected L929 cells from TNFalpha-mediated cytotoxicity, while low expression (about 4 fg/cell/48 h) had no effect. However, clusterin expression did not provide transfected L929 cells with protection against death mediated by colchicine, staurosporine or azide. High level expression of clusterin in transfected L929 cells also potentiated the cytotoxicity of TGFbeta. It had previously been shown that exposure of L929 cells to TGFbeta provides protection against TNFalpha. We showed that this protective effect is not additive to that of clusterin expression. One interpretation of this data is that it suggests that clusterin and TGFbeta may act via a common mechanism to provide protection against the cytotoxicity of TNFalpha. Our results indicate that an intracellular action of clusterin protein is responsible for protection against TNFalpha cytotoxicity. Exposure to TNFalpha induces an increase in the level of cell-associated clusterin and specifically in the level of a novel clusterin molecule, which when analyzed under reducing conditions by SDS/PAGE and immunoblotting appears as two closely spaced bands at about 36 and 38.5 kDa. When analyzed under the same conditions, the normal form of intracellular clusterin, which is present with or without exposure to TNFalpha, appears as two poorly resolved bands at about 43-45 kDa. Since the novel form of clusterin is also expressed in cells exposed to TGFbeta, colchicine, staurosporine, and azide, it may result from toxin-induced disruption of processes of normal cellular protein production.

Animals↗
PubMed1992

Intrarenal distribution of clusterin following reduction of renal mass.

Clusterin is a multifunctional protein isolated from a number of tissues in several different species. In a variety of renal diseases, clusterin appears in the glomerulus and tubules in association with the membrane attack complex of complement. It is also transiently expressed after several forms of acute renal injury. In this study, we examined the expression and intrarenal distribution of clusterin following subtotal renal ablation. Male rats were subjected to either 1-1/3 nephrectomy (1-1/3 NX), uninephrectomy (UNX) or sham operation (SHAM). Two weeks after surgery, clusterin mRNA was elevated in the 1-1/3 NX group (1-1/3 NX: 1215 +/- 88; UNX: 208 +/- 11; SHAM: 207 +/- 19 OD units; P less than 0.001). Clusterin mRNA increased between 3 and 24 hours after 1-1/3 NX, plateaued, and remained elevated for at least seven weeks. The increased clusterin mRNA in 1-1/3 NX was localized to the tissue adjacent to the infarctive scar (scar 858 +/- 173 vs. non-scar 98 +/- 27 OD units; P less than 0.001). Clusterin protein followed a similar pattern of localization, being increased in most tubules and some peritubular capillaries in the peri-infarct zone. Only occasional tubules were positive for clusterin in the renal tissue distant from the scar or in the kidneys of sham operated rats. Co-localization of clusterin and C5b-9 was not detected. Evidence for apoptosis was found in the peri-infarct zone but not elsewhere in 1-1/3 NX kidney or in the normal kidney following sham operation. Infarction of 1/3 of the left kidney without contralateral nephrectomy, a maneuver which eliminates the compensatory growth, and uremia seen with 1-1/3 NX still resulted in increased clusterin mRNA in the infarcted left kidney compared to the intact right kidney (LK: 790 +/- 112 vs. RK: 128 +/- 25 OD units; P less than 0.001), although the amount of clusterin mRNA was less than that found following 1-1/3 NX. In conclusion, persistently increased clusterin mRNA and protein was seen in the peri-infarct zone following 1-1/3 NX. This increased expression of clusterin may be playing a role in the ischemia-related apoptosis present in the scar-adjacent tissue.

Animals↗
PubMed1994

Expression of clusterin in human renal diseases.

Clusterin, a glycoprotein with potent cohesive properties, is induced in a wide variety of acute and chronic experimental renal diseases. The purpose of this study was to examine clusterin expression in human renal diseases. Clusterin immunostaining was examined in nephrectomy specimens from patients with autosomal-dominant polycystic kidney disease (N = 5), autosomal-recessive polycystic kidney disease (N = 3), multilocular cyst of the kidney (N = 2), renal hypoplasia/dysplasia (N = 7), Wilms' tumor (nephroblastoma) (N = 6), renal cell carcinoma (N = 9), and acute and/or chronic renal transplant rejection (N = 15). No clusterin staining was detected in normal renal tissue distant from renal cell carcinomas. Increased expression of clusterin was found in epithelial cells lining cysts in all of the cystic disorders studied. Clusterin expression was found in some immature tubules in hypoplastic/dysplastic kidneys and in tubules of rejected renal allografts, but was not a prominent finding in renal neoplasms, although some renal cell carcinomas expressed clusterin in a focal manner. Common features of clusterin induction included exclusively epithelial production of clusterin in cysts, immature nephrons, and injured tubules, heterogeneity of clusterin expression, with only some tubules and/or cysts in a given area staining for clusterin, and uniform clusterin staining of epithelial cells in a given tubule or cyst in most cases. Based on its cohesive properties, we speculate clusterin functions to maintain cell-cell and cell-substratum interactions which become perturbed in the setting of renal injury and cystic diseases.

Adolescent↗
PubMed2002

Depletion of clusterin in renal diseases causing nephrotic syndrome.

BACKGROUND: Clusterin is a lipoprotein that has anti-complement effects in membranous nephropathy (MN). In focal segmental glomerulosclerosis (FSGS), it inhibits permeability plasma factor activity and could influence proteinuria. Moreover, with aging, knockout mice for clusterin develop a progressive glomerulopathy with sclerosis. METHODS: Since little is known about clusterin metabolism in humans, we determined clusterin levels and composition in the sera and urine of 23 patients with MN, 25 with FSGS and 23 with steroid-responsive nephrotic syndrome (NS). Renal localization was evaluated by immunofluorescence and morphometry. RESULTS: Serum clusterin was markedly reduced in active MN, in FSGS and in children with NS compared to controls; after stable remission of proteinuria, nearly normal levels were restored. Among various biochemical variables, serum clusterin was inversely correlated with hypercholesterolemia. Urinary clusterin, representing a 0.01 fraction of serum, was higher in the urine from normal subjects and FSGS patients in remission with proteinuric MN, FSGS and idiopathic NS; clusterin was inversely correlated with proteinuria. In all cases, urinary and serum clusterin was composed of the same 80 kD isoforms. Finally, a decrease in focal segmental or global clusterin staining was found in FSGS glomeruli, especially in areas of sclerosis. Instead, in MN an overall increment of staining was observed that ranged from mild/focal to very intense/diffuse. CONCLUSIONS: The overall pool of clusterin is reduced in glomerular diseases causing nephrotic syndrome, with hypercholesterolemia appearing as the unifying feature. Depletion of clusterin should negatively affect the clinical outcome in nephrotic patients and efforts should be aimed at normalizing clusterin overall pool.

Adolescent↗
PubMed2005

Antisense oligodeoxynucleotide therapy targeting clusterin gene for prostate cancer: Vancouver experience from discovery to clinic.

BACKGROUND: The objective of this study was to review our experience in the development of antisense (AS) oligodeoxynucleotide (ODN) therapy for prostate cancer targeting antiapoptotic gene, clusterin. METHODS: We initially summarized our data demonstrating that clusterin could be an optimal therapeutic target for prostate cancer, then presented the process of developing AS ODN therapy using several preclinical animal models. Finally, the preliminary data of the recently completed phase I clinical trial using AS clusterin ODN as well as the future prospects of this therapy are discussed. RESULTS: Expression of clusterin was highly up-regulated after androgen withdrawal and during progression to androgen-independence, but low or absent in untreated tissues in both prostate cancer animal model systems and human clinical specimens. Introduction of the clusterin gene into human prostate cancer cells confers resistance to several therapeutic stimuli, including androgen ablation, chemotherapy and radiation. AS ODN targeting the translation initiation site of the clusterin gene markedly inhibited clusterin expression in prostate cancer cells in a dose-dependent and sequence-specific manner. Systemic treatment with AS clusterin ODN enhanced the effects of several conventional therapies through the effective induction of apoptosis in prostate cancer xenograft models. Based on these findings, a phase I clinical trial was completed using AS clusterin ODN incorporating 2'-O-(2-methoxy)ethyl-gapmer backbone (OGX-011), showing up to 90% suppression of clusterin in prostate cancer. CONCLUSIONS: The data described above identified clusterin as an antiapoptotic gene up-regulated in an adaptive cell survival manner following various cell death triggers that helps confer a phenotype resistant to therapeutic stimuli. Inhibition of clusterin expression using AS ODN technology enhances apoptosis induced by several conventional treatments, resulting in the delay of AI progression and improved survival. Clinical trials using AS ODN confirm potent suppression of clusterin expression and phase II studies will begin in early 2005.

Clinical Trials as Topic↗
PubMed2000

Clusterin protein diversity in the primate eye.

PURPOSE: The clusterin gene encodes a multi-functional protein that has been identified in different tissues, including a number of different eye tissues, primarily in the mouse and to a much lesser extent in humans. Clusterin has been implicated in a number of cellular processes such as lipid transport, membrane integrity, apoptosis, and neurodegeneration, all of which could be important to the biology of the eye. In the current communication, we provide data that confirms the expression of clusterin in a number of different human eye tissues and establishes the expression profile of this gene in monkey derived eye tissues. The issue that we sought to examine is whether a broad profile of clusterin expression in the eye is consistent in primates (monkey and human). METHODS: The majority of our study was done using monkey eye tissues. Where possible, we have used human tissues in order to confirm published findings. Northern and western analysis was performed using tissues derived from monkey eyes. In situ hybridization and immunochemistry were carried out on human eye sections. RESULTS: Clusterin mRNA is expressed in primate lens, cornea, limbus, sclera, orbital muscle, ciliary body, retina, RPE/choroid, and RPE cells in culture. Western analysis revealed that two major groups of clusterin exist in the eye, a high molecular weight group (>100 kDa) and a second group consisting of at least five clusterin species that are all approximately 80 kDa. Analysis of conditioned media from RPE cells cultured on permeable supports suggests that different forms of clusterin display alternative patterns of secretion. CONCLUSIONS: Clusterin is expressed in a broad range of eye tissues in both human and monkey, suggesting that this is a characteristic feature in primates. We demonstrate for the first time that a diverse number of clusterin isoforms were observed in monkey eye tissues by western analysis. Meanwhile, the molecular size of clusterin mRNA detected in the array of tissues are identical in size, suggesting that the nature of the diversity in clusterin forms is due to post-translational modifications. In addition, new insights were made in defining clusterin expression in ciliary body, cornea, and the retinal pigment epithelium.

Animals↗
PubMed1999

A reexamination of the role of clusterin as a complement regulator.

Clusterin is a highly conserved glycoprotein which has been proposed to protect host cells against complement-mediated cytolysis. We tested the hypothesis that clusterin is a complement regulator using erythrocytes and cells which had been stably transfected with a membrane-anchored form of clusterin as targets for complement-mediated cytolysis. Clusterin gave dose-dependent protection of antibody-coated sheep erythrocytes against complement-mediated lysis by diluted normal human serum. There was a linear relationship between the concentration of clusterin giving 50% protection and the concentration of serum; extrapolation of this to the case of undiluted human serum showed that a clusterin concentration at least two orders of magnitude greater than its physiological plasma concentration would be needed to confer protection against complement-mediated cytolysis under physiological conditions. Physiological concentrations of clusterin did not protect rabbit erythrocytes against alternative complement pathway-mediated lysis using dilute human serum. Exogenous clusterin had no effect on lysis of human erythrocytes triggered by the addition of inulin to autologous human serum. Induction of cell-surface clusterin expression by L929 (murine fibroblast) cells which had been stably transfected with cDNA for human clusterin linked to DNA coding for the 44 C-terminal amino acid residues of CD55 did not protect the cells against complement-mediated lysis by either normal or clusterin-depleted human serum. These data suggest that clusterin may not be a physiologically relevant regulator of complement activation.

Animals↗
PubMed2001

Clusterin expression during regeneration of pancreatic islet cells in streptozotocin-induced diabetic rats.

AIMS/HYPOTHESIS: Beta-cell regeneration has been reported after islet injury in an animal model for diabetes. Recently, we showed up-regulation of clusterin after islet injury and suggested that clusterin might be involved in cytoprotection and in the regeneration of islet cells. The aim of this study was to investigate the correlation of clusterin expression with islet regeneration and its effect on islet cell replication. METHODS: Streptozotocin was administrated to rats to induce various types of diabetes. Islet regeneration and clusterin expression were examined after islet injuries. Clusterin cDNA was transfected to MIN6 cells and their proliferation activity was measured by a [3H]thymidine-incorporation assay. RESULTS: A diabetogenic dose of streptozotocin injected in rats provoked an immediate degeneration of beta cells. In this model, islets showed increased clusterin expression with extensive proliferation of alpha cells but showed poor beta-cell replication. A subdiabetogenic dose of streptozotocin, however, led to the proliferation of beta cells with clusterin up-regulation. In streptozotocin-treated neonatal rats, up-regulation of clusterin was noted during beta-cell proliferation. In all experimental models, clusterin was expressed in alpha cells in close correlation with islet cell proliferation, higher transcription of insulin mRNA and MAPKs activation. Cell replication was increased by 31 % in the MIN6 cells transfected by the clusterin cDNA. CONCLUSION/INTERPRETATION: Up-regulation of clusterin in alpha cells might induce beta-cell proliferation and thus restore their population after islet injury. We suggest that clusterin could be considered as a growth factor-like molecule stimulating islet-cell proliferation by paracrine action.

Aging↗
PubMed2005

Use of antisense oligonucleotides targeting the cytoprotective gene, clusterin, to enhance androgen- and chemo-sensitivity in prostate cancer.

The discovery and targeting of genes mediating androgen-independence may lead to the development of novel therapies that delay progression of hormone refractory prostate cancer (HRPC). Clusterin is a stress-associated cell survival gene that increases after androgen ablation. Here, we review clusterin's functional role in apoptosis and the use of antisense oligonucleotides (ASOs) against clusterin to enhance apoptosis in prostate cancer models. Immunostaining of tissue microarrays constructed from untreated and post-hormone treated radical prostatectomy specimens confirm that clusterin is highly expressed in virtually all HRPC cells, 80% of prostate cancer cells after neoadjuvant hormone therapy, but is low or absent (<20%) in untreated specimens. Overexpression of clusterin in LNCaP cells confers resistance to both androgen ablation and chemotherapy. Clusterin ASOs reduced clusterin levels in a dose-dependent and sequence-specific manner. Adjuvant treatment with murine clusterin ASOs after castration of mice bearing Shionogi tumors decreased clusterin levels, accelerated apoptotic tumor regression, and significantly delayed the recurrence of androgen-independent tumors. A human clusterin ASO targeting the translation initiation site and incorporating MOE-gapmer backbone (OGX-011) synergistically enhanced the cytotoxic effects of paclitaxel in human xenografts of prostate, renal cell, bladder, and lung cancer. Clusterin, is an anti-apoptosis protein upregulated in an adaptive cell survival manner by androgen ablation and chemotherapy that confers resistance to various cell death triggers. Suppression of clusterin levels using ASOs enhances cell death following treatment with androgen ablation, radiation, and chemotherapy.

Androgens↗
PubMed1998

Clusterin overexpression in rat pancreas during the acute phase of pancreatitis and pancreatic development.

Molecular mechanisms associated with apoptosis in pancreas remain largely unknown. Clusterin mRNA is induced in several tissues in response to most apoptotic stimuli. In these tissues, clusterin has an antiapoptotic activity. The aim of this work was to test whether clusterin, which is not expressed in normal pancreas, was induced in pancreas during pancreatitis and pancreatic development. Clusterin mRNA levels were strongly increased 6 h after pancreatitis induction. Maximal expression happened between 24-48 h and decreased progressively to undetectable levels at day 5. Clusterin mRNA was expressed with similar intensity in oedematous caerulein-induced pancreatitis and in response to various degrees of necrohaemorrhagic taurocholate-induced pancreatitis, indicating a maximal gene activity in all types of pancreatitis; in situ hybridization showed that the acinar cells and some ducts expressed clusterin mRNA. A single band of about 35-38 kDa was detected by western blot in pancreatic homogenates and in pancreatic juice from rats with acute pancreatitis, but not from control rats. Clusterin mRNA expression was strong in late fetal life and remains high until day 11 post-partum, then decreased progressively with a minimum from 35 to 90 days post-partum. Clusterin mRNA levels were strongly induced in pancreatic acinar AR4-2J cells in response to various apoptotic stimuli (i.e., cycloheximide, staurosporine, ceramide and H2O2) but not with interleukin (IL)-1, IL-4 or IL-6 or heat shock, which do not induce apoptosis in AR4-2J cells. In conclusion, we demonstrated that clusterin is synthesized and released by the pancreas. Its strong expression during acute pancreatitis suggests its involvement in the pancreatic response to injury. Clusterin is also induced during pancreatic development. Because these situations are associated with apoptosis and clusterin was shown to protect against apoptosis, we speculate that clusterin could be involved in the control of acinar cell apoptosis.

Acute Disease↗
PubMed2001

Clusterin expression in adult human normal and osteoarthritic articular cartilage.

OBJECTIVE: To characterize the expression pattern of clusterin in adult human normal and osteoarthritic cartilage. METHODS: Clusterin mRNA expression in adult human normal and osteoarthritic cartilage was investigated by analysis of cDNA libraries, TaqMan quantitative RT-PCR, microarray and in situ hybridization. RESULTS: Sequence analysis of ESTs from adult human normal and osteoarthritic cartilage cDNA libraries demonstrated that the abundance of clusterin in these libraries was equivalent to genes which have been more commonly associated with cartilage. To examine tissue distribution, TaqMan Quantitative PCR analysis was performed using RNA from a panel of individual normal tissues. Clusterin was expressed at significant levels in cartilage, brain, liver, and pancreas. The expression of clusterin mRNA was up-regulated in early osteoarthritic vs normal cartilage when analysed by microarray analysis. Using in situ hybridization, chondrocytes of normal cartilage expressed moderate levels of clusterin. Upper mid-zone chondrocytes in cartilage with early stages of osteoarthritic disease expressed high levels of clusterin mRNA. In advanced osteoarthritic cartilage, the overall expression of clusterin was reduced. CONCLUSION: The induction of clusterin has been associated with a variety of disease states where it appears to provide a cytoprotective effect. The increased expression of clusterin mRNA in the early stages of osteoarthritis (OA) may reflect an attempt by the chondrocytes to protect and repair the tissue. In contrast, the decrease in clusterin mRNA in the advanced osteoarthritic cartilage accompanies the final degenerative stages of the disease. An understanding of the expression of clusterin in osteoarthritis may allow consideration of this protein as a marker for cartilage changes in this chronic degenerative condition.

Adult↗
PubMed2005

Expression of the secreted form of clusterin protein in renal cell carcinoma as a predictor of disease extension.

OBJECTIVES: To evaluate the significance of clusterin expression in surgically resected renal cell carcinoma (RCC) specimens. PATIENTS AND METHODS: Normal kidney and RCC specimens were obtained from 131 patients who had radical surgery. The expression of clusterin protein was analysed by immunohistochemical staining with an antibody recognizing all isoforms of clusterin. Cell proliferative activities and apoptotic features in these specimens were investigated using Ki-67 immunostaining and the terminal deoxynucleotidyl transferase mediated dUTP nick-end labelling assay, respectively. Findings were evaluated in relation to several clinicopathological factors. RESULTS: There were various levels of clusterin expression in 128 of the 131 RCC specimens, while 37 of 131 normal kidney tissues (28.2%) had no clusterin staining. Clusterin protein was present in the cytoplasm of both normal and cancer cells, but there was no nuclear staining identified in either type of cell. The expression level of clusterin protein in RCC tissues was significantly related to tumour stage and grade, but not to age, gender or histological cell type. Cell proliferative activity in RCC specimens was significantly associated with clusterin expression, while the apoptotic index was inversely related to clusterin expression. Furthermore, recurrence-free survival in patients with strong clusterin expression was significantly lower than that in those with weak expression. CONCLUSIONS: These findings suggest that the secreted form of clusterin may be involved in the progression of RCC, and that overexpression of clusterin could be a useful prognostic variable after radical surgery in patients with RCC.

Adult↗
PubMed1992

Distinct sites of production and deposition of the putative cell death marker clusterin in the human thymus.

Clusterin is a multifunctional protein endowed with cell-aggregating, complement-inhibitory, and lipid-binding properties. Since several studies have demonstrated highly increased clusterin gene expression in epithelial and nervous tissues regressing as a consequence of tissue involution and apoptotic cell death, clusterin is also considered as a specific marker of dying cells. To determine whether clusterin expression is also upregulated during thymocyte death occurring during the negative selection process we analyzed the cellular distribution of clusterin mRNA and protein by in situ hybridization and immunocytochemistry in the human thymus. We observed that the expression of clusterin mRNA was confined to cells present in the thymic medulla, concentrated mainly around Hassal's bodies. Immunostaining of adjacent sections with antikeratin Ab revealed that cells containing clusterin mRNA were predominantly epithelial. By contrast no clusterin mRNA was found in thymocytes by in situ hybridization and Northern blot analysis of total RNA from purified thymocyte populations. Clusterin protein colocalized with the membrane attack complex of complement and vitronectin in the center of the largest Hassal's bodies, but was not detectable by immunocytochemistry in or at the surface of epithelial cells. Our results demonstrate that clusterin gene expression does not take place in apoptotic thymocytes, and therefore that clusterin synthesis by the dying cell is probably not a prerequisite to its death. However, synthesis of clusterin by medullary epithelial cells may be related to their terminal differentiation, and, furthermore, its presence in Hassal's bodies raises the possibility that the secreted protein is involved in the disposal of cell debris resulting from thymocyte apoptosis.

Apoptosis↗
PubMed1995

Clusterin promotes the aggregation and adhesion of renal porcine epithelial cells.

The function of clusterin, a heterodimeric glycoprotein markedly induced in renal and other organ injuries, is unclear. Since renal injury is accompanied by alterations in cell attachment, it is possible that clusterin functions to promote cell-cell and cell-substratum interactions. In this study, a single cell suspension of renal epithelial (LLC-PK1) cells was treated with purified human clusterin, resulting in time- and dose-dependent cell aggregation. Electron microscopy of the cell aggregates demonstrated cell junction and lumen formation. To determine the effect of clusterin on cell adhesion, tissue culture plates were coated with clusterin, fibronectin, PBS, or albumin. Clusterin and fibronectin promoted cell adhesion to the same extent. The adhesion to clusterin was dose dependent and specific, as a monoclonal antibody against clusterin inhibited cell adhesion to clusterin but not fibronectin. Perterbations of the cytoskeleton may underlie the alterations in cell attachment which occur in renal injury. Induction of clusterin mRNA was seen after disruption of both microtubules and microfilaments and after inhibition of cell-substratum interactions. In conclusion, clusterin is a potent renal epithelial cell aggregation and adhesion molecule. We speculate that clusterin functions to promote cell-cell and cell-substratum interactions which are perturbed in the setting of renal injury, thereby preserving the integrity of the renal epithelial barrier.

Animals↗
PubMed1992

Clusterin production in the obstructed rabbit kidney: correlations with loss of renal function.

Clusterin, a protein associated with cell death, has been suggested as a marker of renal injury. Correlation of clusterin gene expression with changes in renal function and quantitative measurement of clusterin protein levels after ureteral obstruction have not been previously reported. With unilateral ureteral obstruction in rabbits as the experimental model, the time course of alterations in renal function, clusterin mRNA accumulation, and concentrations of clusterin protein in serum, urine, and renal tissue were investigated. RBF, GFR, and renal concentrating ability (percent sodium reabsorption and urine osmolarity) all decreased (P < 0.05) in the obstructed kidney from control values within 1 day of ureteral obstruction. Clusterin mRNA levels started to rise in the ipsilateral kidney within 12 h of ureteral obstruction and increased up to 10-fold above control levels after 3 days of obstruction. Hybridization histochemistry showed that clusterin mRNA was initially detectable in collecting ducts and distal tubules within 12 h of ureteral obstruction. After 7 days of obstruction, increased accumulation of clusterin mRNA was also detectable in proximal tubular epithelial cells. Clusterin gene expression remained elevated in collecting ducts after 60 days of obstruction. Clusterin expression in the contralateral kidney was increased twofold over control values after 12 h of obstruction. No increase in clusterin mRNA accumulation was detectable after 24 h in the contralateral kidney. Total clusterin protein in the obstructed kidney increased from 0.59 +/- 0.66 (mean +/- 1 SD) to 2.5 +/- 1.3 micrograms after 7 days of ureteral obstruction (P < 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗