Copyright ©The Histochemical Society, Inc.

Tensin Is Expressed in Glomerular Mesangial Cells and Is Related to Their Attachment to Surrounding Extracellular Matrix

Hisatsugu Takahara, Isao Shirato, Katsuhiko Asanuma, Michifumi Yamashita, Yukihiko Takeda and Yasuhiko Tomino

Division of Nephrology, Department of Internal Medicine, Juntendo University School of Medicine, Tokyo, Japan

Correspondence to: Yasuhiko Tomino, Div. of Nephrology, Dept. of Internal Medicine, Juntendo U. School of Medicine, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113-8421, Japan. E-mail: yasu{at}med.juntendo.ac.jp


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Glomerular expression of tensin was immunohistochemically studied in normal and diseased rat kidneys to determine whether tensin might be related to specific binding in individual glomerular cells. Normal rat kidneys displayed an intense immunofluorescence reaction for tensin along the basal aspects of proximal and distal tubule cells and parietal epithelial cells of Bowman's capsules. In glomeruli, a positive reaction for tensin was detected only in the mesangial areas. Immunoelectron microscopy revealed a positive reaction in the mesangial cell (MC) processes. RT-PCR and immunoprecipitation demonstrated mRNA and protein levels of tensin in cultured rat MCs. Mesangial tensin expression was decreased when the mesangium was injured by Habu snake venom. During the regenerative process after mesangiolysis, tensin expression was not detected in early-phase proliferating MCs that did not have extracellular matrix (ECM). The expression of tensin recovered in late-phase proliferating MCs, which became attached to regenerated ECM. It appears that tensin is related to MC attachment to surrounding ECM, which suggests that signal transduction regulated by tensin may be related to a specific mechanism of MC matrix regeneration. Furthermore, tensin can act as a marker for rat MCs because the expression of tensin was detected only in MCs in glomeruli. (J Histochem Cytochem 52:683–691, 2004)

Key Words: tensin • mesangial cell • mesangial cell marker • Habu snake venom nephritis


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THE GLOMERULAR MESANGIUM occupies the central region of the glomerular lobule to which the glomerular capillaries are affixed (Kriz et al. 1990aGo,bGo). The mesangial cell (MC) has several functions, such as maintenance of glomerular capillary architecture (Sakai and Kriz 1987Go; Lemley et al. 1992Go), phagocytosis of macromolecules, and immune complexes (Schlondorff and Mori 1990Go; Hughes et al. 1997Go; Cortes–Hernandez et al. 2002Go), and production of bioactive substances (Busiek et al. 1995Go; Diaz–Cazorla et al. 1999Go; Rovin et al. 2001Go). MC–matrix interactions at the MC processes may play an important role in MC function (Drenckhahn et al. 1990Go; Gauer et al. 1997Go). In the cell processes, actin microfilament bundles connect to the glomerular basement membrane (GBM) through the interposition of integrin transmembrane protein (Drenckhahn et al. 1990Go). In such cell–matrix adhesion sites, several actin-associated proteins, including {alpha}-actinin, talin, and vinculin, have been identified (Drenckhahn and Franke 1988Go; Drenckhahn et al. 1990Go; Prols et al. 1999Go). These proteins bind actin and integrin, and some of them are related to signal transduction (Gauer et al. 1997Go; Prols et al. 1999Go). It is generally considered that {alpha}8 integrin is specific for MCs (Hartner et al. 1999Go; Bieritz et al. 2003Go). However, the actin-associated protein specific for MC remains unknown.

Tensin is an actin-associated 220-kD phosphoprotein located between actin and integrin (Lo et al. 1994aGo; Chuang et al. 1995Go; Chen et al. 2000Go). The protein contains a functional Src homology 2 (SH2) domain (Davis et al. 1991Go), which is able to interact with certain tyrosine phosphorylated proteins (Salgia et al. 1995Go). In addition, it binds to barbed ends of actin filaments and is able to crosslink with actin filaments (Lo et al. 1994bGo; Chuang et al. 1995Go). Because of these properties, tensin has a dual role in cytoskeletal architecture and signal transduction (Lo et al. 1994cGo). A previous study using tensin-null mice showed that tensin was expressed in the tubule epithelial cells and was related to focal adhesion of these cells to the tubular basement membrane (Lo et al. 1997Go). However, expression of tensin in the glomeruli has not been examined.

In the present study, the localization of tensin was examined by immunohistochemistry (IHC) in rat kidneys, and selective localization of tensin in the glomerular MCs was identified. In addition, to explore the putative role of tensin in renal disease with mesangiolysis, the expression of tensin was analyzed in Habu snake venom (HSV) nephritis. Pathological lesions of HSV nephritis induced in Sprague–Dawley (SD) rats were characterized by mesangiolysis in the initial phase and MC proliferation in the late phase (Bradfield et al. 1977Go; Cattell and Bradfield 1977Go; Barners et al. 1994aGo).


    Materials and Methods
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 Materials and Methods
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Animals
All animal experiments were conducted in accordance with the National Research Council Guidelines for the Care and Use of Laboratory Animals. Adult male SD rats weighing ~200 g were obtained from Japan Charles River Laboratories (Kanagawa, Japan).

Induction of Glomerular Disease Models
HSV nephritis was induced in adult SD rats by a single IV injection of HSV at 4.0 mg/kg body weight (Wako Pure Chemical Industries; Osaka, Japan). In addition to a control group (no injection), two experimental groups with animals sacrificed at 5 days or 10 days after the HSV injection were studied.

Fixation of Kidneys
After the rats were anesthetized with sodium pentobarbital (100 mg/kg body wt), they were perfused via the abdominal aorta with 4% paraformaldehyde (PFA) in PBS, pH 7.2, at 200 mmHg for 5 min. After perfusion of PFA, the left kidney was cut in half, dipped in 10% formaldehyde at 4C, and processed and embedded in paraffin for light microscopy. For electron microscopy, one half of the right kidney was cut into several pieces and fixed with 2% glutaraldehyde (GA) in 0.1 mol/liter phosphate buffer (PB) at 4C for 2 hr. After the tissues were washed in PB, they were postfixed with 2% aqueous osmium tetroxide at room temperature (RT) for 2 hr. Then they were dehydrated in a graded ethanol series at RT and finally embedded in Epon 812 by standard procedures. After perfusion of PFA, the right kidney was perfused with cryoprotectant 20% sucrose in PBS at 200 mmHg for 5 min. The kidney was removed and dipped in 20% sucrose solution at 4C for 2 hr. Finally, the kidney was cut into several pieces and frozen in melting isopentane cooled by liquid nitrogen to –160C for IHC.

Light and Electron Microscopy
For light microscopy, the paraffin-embedded tissues were cut into 3-µm-thick sections, mounted and stained with periodic acid–Schiff (PAS) by standard procedures. The sections were examined with an Olympus light microscope (Olympus; Tokyo, Japan).

For electron microscopy, the tissues were cut with a regular diamond knife into semi- and ultrathin sections. Semi-thin sections (1 µm thick) stained with 1% toluidine blue were used for light microscopy. Ultrathin sections (silver to gold) were collected on 100-mesh copper grids and double stained with 4% uranyl acetate and lead citrate. The sections were then examined with a Hitachi 7100 transmission electron microscope (Hitachi High Technologies; Tokyo, Japan) operated at 75 kV.

Immunohistochemistry
Frozen renal sections were cut into 3-µm-thick sections using the cryostat. As the primary antibody (Ab), a mouse monoclonal anti-tensin antibody (BD Transduction Laboratories; BD Biosciences, San Jose, CA) was used (Dugina et al. 2001Go; Kook et al. 2003Go). The second Ab was FITC-conjugated goat anti-mouse IgG antiserum (Cappel; ICN Pharmaceuticals., Aurora, OH). After the sections were blocked with PBS containing 2% bovine serum albumin, 2% fetal calf serum, and 0.2% fish gelatin for 15 min, they were incubated with primary Ab diluted 1:100 in blocking solution at room temperature for 60 min. The sections were then incubated with secondary Ab diluted 1:100 in blocking solution with 10% normal rat serum at RT for 30 min. After immunostaining, the sections were examined with a Leica fluorescence microscope (Leica Microsystems; Tokyo, Japan).

For immunoelectron microscopy, the frozen renal sections were cut into 3-µm-thick sections with a cryostat. After the sections were blocked with blocking solution for 15 min, they were incubated with primary Ab diluted 1:100 in blocking solution at 4C overnight. The sections were incubated with biotinylated goat anti-mouse IgG at 4C for 5 hr and were then incubated with avidin–peroxidase complexes from the Vectastain Elite ABC Kit (Vector Laboratories; Burlingame, CA) at RT for 30 min, followed by incubation with the peroxidase substrate solution diaminobenzidine in PBS for a few minutes. The sections were fixed with 2% GA at 4C for 60 min. They were then dehydrated in a graded series of ethanol at RT and finally embedded in Epon 812 by standard procedures.

For immunostaining of rat MCs (described below), two types of cells maintained in normal medium or FCS-free medium for 5 days were used. Primary rat MCs were placed on collagen type I-coated glass coverslips (Iwaki Glass; Tokyo, Japan) and cultured continuously with normal medium or FCS-free medium for 12 hr at 37C. After the cells were washed with PBS, they were fixed with 3% PFA containing 0.5% Triton X-100 (Sigma; St Louis, MO) for 5 min and then fixed with 3% PFA for 20 min. After the cells were incubated with blocking solution for 30 min, they were incubated with anti-tensin Ab diluted 1:100 in blocking solution at RT for 60 min. The cells were then incubated with AlexaFluor 488 goat anti-mouse IgG Ab (Molecular Probes; Eugene, OR) diluted 1:200 in blocking solution at RT for 60 min. After immunostaining, the cells were examined with a confocal microscope (Japan Bio-Rad Laboratories; Tokyo, Japan).

Preparation of Rat Glomeruli and Culture of Primary Rat MCs
After the rats were anesthetized, the kidneys were removed for RNA and protein extraction. Glomeruli were isolated by differential sieving methods (Striker et al. 1987Go). The glomeruli were incubated on type I collagen-coated 10-cm dishes to obtain MCs. Primary rat MCs were maintained in Dulbecco's modified Eagle's medium (Sigma) supplemented with 5% FCS, 100 U/ml penicillin G, and 100 mg/ml streptomycin at 37C.

RNA Preparation and RT-PCR in Rat MCs and Kidney Cortex
Rat MCs (1.2 x 106) were incubated on type I collagen-coated 10-cm dishes. Total RNA of rat MCs and kidney cortex were extracted with an RNeasy Mini Kit (QIAGEN; Hilden, Germany) for RNA preparation. One microgram of total RNA was reverse-transcribed using oligo (dT) primers (Life Technologies; Rockville, MD) and reverse transcriptase (Superscript II; Life Technologies). The single-strand cDNA product was denatured and amplified in the Gene Amp PCR System 9600 (Perkin–Elmer; Norwalk, CT) with two primers under the following conditions: 1 min at 95C; then 30 sec at 94C, 30 sec 54C and 1 min 72C for 35 cycles; and finally 72C 10 min and 4C. The cDNAs were amplified using various primers specific only for cDNA sequences of rat tensin (forward 5'-ACATTACCCCCTCAACACTG-3' and reverse 5'-GTTCTGGATTCAAGGTGGTC-3'). The amplified products were confirmed by sequencing. The amplified products were electrophoresed with a 2% agarose gel, stained with ethidium bromide, and photographed under a UV lamp.

Immunoprecipitation of Rat MCs and Kidney Cortex
Rat MCs and kidney cortex were lysed in 1% NP-40 buffer [1% NP-40 = 300 mM NaCl, 100 mM Tris-HCl (pH 7.5), 0.1% sodium dodecyl sulfate (SDS), 2 mM ethylenediaminetetraacetate] containing proteinase inhibitors (1 mM phenylmethylsulfonyl fluoride, 1 mM pepstatin A, 1 mM E-64C) on ice. Insoluble tissue lysates were removed by centrifugation at 15,000 x g for 5 min. The lysates were subjected to immunoprecipitation with a mouse monoclonal anti-tensin antibody and protein G-PLUS-Agarose (Santa Cruz Biotechnology; Santa Cruz, CA) at 4C overnight. The immunoprecipitates were collected on protein G-PLUS-Agarose, washed with NP-40 buffer, and then extracted from the beads by boiling in SDS sample buffer. They were analyzed by immunoblot analyses. Immunoblotting analyses were carried out according to standard protocols (Laemmli 1970Go; Towbin et al. 1979Go) and visualized by chemiluminescent methods with ECL plus immunoblot detection kits (Amersham Pharmacia Biotech; Poole, UK).


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Immunofluorescence
In the cortex of normal rat kidney, a positive reaction for tensin was detected in tubuli, Bowman's capsules, and glomeruli (Figure 1A) . The expression of tensin in tubuli displayed a continuous linear fluorescence at the basal aspects, but the intensity of the immunoreactivity was variable among tubule segments. Proximal convoluted tubuli were more intensely stained than other segments. In Bowman's capsules, the positive reaction was also detected at the basal aspects of parietal epithelial cells. The distribution of tensin was similar to that of actin fiber bundles in the cells. When the cells were cut vertically, linear immunoreactivity was detected in the cells. When the cells were cut obliquely, the immunoreactivity appeared as several short parallel lines. In glomeruli, a positive reaction for tensin was detected in the mesangium (Figure 1B). The intensity of the positive reaction was almost identical in each mesangial area of individual glomeruli. Other components in the cortex, i.e., blood vessels, interstitial cells and glomerular capillary walls, did not react with tensin.



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Figure 1

Immunofluorescence of normal rat kidney for tensin. (A) Low-magnification view of cortex demonstrates localization of tensin in tubuli, Bowman's capsules, and glomeruli. In tubuli and Bowman's capsules, expression of tensin is detected linearly along basal aspects of the cell. (B) In a section of glomerulus, expression of tensin is observed only in the mesangial areas. Bars: A = 50 µm; B = 20 µm.

 
Immunoelectron Microscopy
The presence of tensin in tubule epithelial cells and parietal epithelial cells of Bowman's capsules was clearly observed by immunoelectron microscopy. In each cell, immunoreactive products were localized at the inner aspects of the basal cell membranes attached to individual basement membranes (Figures 2A–2C) . In the mesangium, immunoreactivity for tensin was localized in MCs (Figure 2D). Most of the immunoreactivity was localized in processes of MCs. Among individual processes, tongue-like processes extending toward the mesangial angles of the GBM exhibited the strongest reactivity (Figure 2E).



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Figure 2

Immunoelectron microscopy of normal rat kidney for tensin. (A) In the tubuli, a positive reaction for tensin is detected in the basal aspect of both proximal (P) and distal (D) tubule epithelial cells. (B) In a section of a proximal tubule epithelial cell, tensin is localized at the inner aspect of the tips of the epithelial cell invaginations where the epithelial cell contacts the tubular basement membrane (TBM). (C) In the parietal epithelial cells of Bowman's capsule, a positive reaction for tensin is detected in the basal aspects of the cell. (D) In a cross-section through the mesangial area, a positive reaction for tensin is detected in the MC processes (arrows), which attach to the mesangial matrix and GBM. (E) Tensin is expressed most strongly in the tongue-like process of MC (arrows) at the mesangial angle which attaches to the GBM. Bars: A = 2 µm; B–D = 1 µm. En, endothelial cell; Po, podocyte.

 
RT-PCR and Immunoprecipitation
Using RT-PCR, a single band of 324 bp for tensin was detected in cultured rat MCs and kidney cortex (Figure 3A) . Immunoprecipitation demonstrated a single immunoreactive band at a position of 220 kD in extracts of cultured rat MCs and kidney cortex (Figure 3B).



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Figure 3

(A) RT-PCR of cultured MCs from normal rat kidney. A single band of 324 bp for tensin is detected. (B) Immunoprecipitation of cultured MCs from normal rat kidney. A single immunoreactive band of 220 kD is detected. Lane 1, mesangial cells; Lane 2, kidney cortex.

 
Cultured Mesangial Cells
In MCs cultured in normal medium, tensin was expressed in the ends of the actin filament where the cells were attached to the substrate (Figure 4A) . Tensin was expressed as elongated or dot-like structures. MCs cultured in FCS-free medium also expressed tensin in the ends of the actin filament (Figure 4B). However, it appeared that the intensity was slightly increased and the elongated structures changed to broader structures.



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Figure 4

(A) Cultured MCs from normal medium. Tensin is expressed at the end of the actin filament as elongated or dot-like structures. (B) Cultured MCs from FCS-free medium. Tensin is also expressed at the end of the actin filament. Compared with A, the intensity of tensin is slightly increased and the elongated structures changed to broader structures. Bars = 20 µm.

 
Habu Snake Venom Nephritis-induced SD Rat Kidney
The expression of tensin was decreased in the mesangial areas of mesangiolytic glomeruli, although a positive reaction for tensin similar to that in normal rat glomeruli was detected in most glomeruli. At day 5, the early phase of the regenerative process of the mesangium, the positive reaction for tensin was still decreased in mesangial areas (Figure 5A) . However, at day 10, the late phase of the regenerative process, a strong reaction for tensin was observed in mesangial areas (Figure 5B). In both glomeruli, {alpha}-smooth muscle actin ({alpha}-SMA) was detected (data not shown) as previously reported (Barners et al. 1994aGo,bGo). In PAS-counterstained sections of the same IF sections, MC proliferation was observed in mesangial areas of each glomeruli (Figures 5C and 5D).



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Figure 5

Changes in expression of tensin in the regenerative process after mesangiolysis by HSV nephritis on day 5 (A,C,E) and day 10 (B,D,F) glomeruli. In immunofluorescence for tensin, expression disappears in the mesangial area (arrow) at day 5 (A), although strong expression is observed in the mesangial area at day 10 (B). Proliferative MCs are confirmed in the mesangial area of both stages by PAS-counterstained sections (C,D). In electron microscopy, MCs at day 5 are in direct contact with neighboring MCs not via the ECM (E). However, MCs at day 10 contact other MCs via the regenerated ECM (F). Bars: A–D = 15 µm; E,F = 2 µm. En, endothelial cell; Po, podocyte.

 
Proliferative MCs showed a significant difference in both phases by electron microscopy. At day 5, most proliferative MCs did not have surrounding extracellular matrix (ECM) and were in direct contact with neighboring MCs (Figure 5E). Some of the MCs had ECM but most of the ECM was basically immature. At day 10, proliferative MCs were also observed in mesangial areas. However, they had regenerative surrounding ECM and were in contact with other MCs through the mature ECM (Figure 5F).


    Discussion
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Tensin-1 is a 220-kD protein localized in the transmembrane junctions between the ECM and the cytoskeleton (Lo et al. 1994aGo; Chuang et al. 1995Go; Chen et al. 2000Go). A recent study suggests that tensin-2 (170 kD), which has some homology to tensin-1, is also localized in focal adhesions in human tissues (Chen et al. 2002Go). Tensin is particularly interesting in the myriad of focal adhesion proteins because it can bind to actin microfilaments at multiple sites (Lo et al. 1994bGo), enabling tensin both to cap the growing ends of actin filaments and to crosslink actin filaments (Lo et al. 1994bGo; Chuang et al. 1995Go). In addition, tensin is phosphorylated on tyrosine residues (Salgia et al. 1995Go) and contains an SH2 domain (Davis et al. 1991Go), suggesting that tensin might participate in signal transduction cascades relevant to various cellular functions, including cell migration (Chen et al. 2002Go; Chen and Lo 2003Go), differentiation (Hiura et al. 1995Go), regeneration (Ishii and Lo 2001Go), and apoptosis (Kook et al. 2003Go). Several factors, including the ECM, platelet-derived growth factor, thrombin, angiotensin, and oncogenes, induce tensin-mediated signal transduction (Chen et al. 2002Go). The presence of these characteristics in a single molecule suggests that tensin may contribute to MC function.

The present study demonstrates that tensin, in addition to its localization in the tubuli and Bowman's capsules, is specifically expressed in the rat glomerular mesangial areas. Subcellular distribution of tensin examined by immunoelectron microscopy revealed that its protein is mostly localized in MC processes that have well-developed microfilament bundles and are in contact with the perimesangial GBM. Immunoprecipitation and RT-PCR using cultured rat MCs confirmed the presence of a protein of 220-kD molecular mass identical to that of tensin and the expression of mRNA for the protein. Although we did not check the specificity of the immunoreactivity by a preincubation test with a corresponding antigen, these findings strongly suggest that tensin itself is contained in MCs of rat glomeruli.

In this study, the expression of tensin was clearly localized in the MC processes where microfilament bundles terminate in the cell membrane to connect with the surrounding GBM. Among these processes, the most prominent expression of tensin was detected in juxtacapillary tongue-like processes that interconnect with the GBM at the two opposing mesangial angles of a single capillary. The distending forces of the glomerular capillary wall are balanced at the mesangial angles and at the mesangium–GBM interface by a complex anchoring system that connects the GBM to the mesangial processes (Sakai and Kriz 1987Go; Kriz et al. 1990aGo,bGo; Lemley et al. 1992Go). These findings point to the essential involvement of tensin in the mechanical role of MCs, i.e., stabilization of glomerular capillary architecture. On the basis of findings that tensin can bind to actin microfilaments at multiple sites, as described above, it is reasonable to assume that tensin expressed in the MC processes is an important molecule for the contractile and adhesion apparatus of MCs. In addition, a recent study reported that MCs organize the glomerular capillaries by adhering to the G- domain of laminin {alpha}5 in the GBM via integrin {alpha}3ß1 and Lu, a member of the immunoglobulin superfamily (Kikkawa et al., 2003Go). Therefore, we suggest that MCs maintain the glomerular capillary architecture in various ways.

The transient strong expression of tensin in MCs during the recovery phase after mesangiolysis induced by HSV suggests that some specific mechanism of MC regeneration might be regulated by tensin-mediated signal transduction. Background data permitting speculation about specific function of the protein are lacking, but it is noteworthy that expression is detected not in early but in late regenerative processes after HSV injury. The same late appearance is also observed in the deposition of ECM such as fibronectin (Barners et al. 1994bGo,1995Go). As demonstrated in cell culture studies (Gauer et al. 1997Go; Prols et al. 1999Go), MCs and matrix proteins might interact to maintain cellular function, resulting in the recovery of normal structure and function of the mesangium.

Recent studies showed that ECM adhesion formed by cultured fibroblasts can be classified into two major types by molecular and structural criteria, i.e., classical focal contact (FC) and fibrillar adhesion (FA) (Zamir et al. 1999Go; Katz et al. 2000Go). These were characterized by distinct complements of integrins and cytoskeletal anchor molecules. FCs are located mainly at the cell periphery, bound mainly to vitronectin fibrils, and contain relatively high levels of vinculin, paxillin, {alpha}-actinin, talin, focal adhesion kinase, and {alpha}vß3 integrin. On the other hand, FAs are located more centrally in cells, are characteristically elongated or dot-like structures, are rich in tensin and {alpha}5ß1 integrin, and are bound to fibronectin fibrils (Zamir et al. 1999Go; Katz et al. 2000Go). These two types of adhesion appear to differ not only in their shape and molecular composition but also in their functions. FCs cannot translocate smoothly as a result of contractile forces and develop high tension. In contrast, FAs are moved dynamically by actomyosin-driven pulling (Zamir et al. 2000Go; Zamir and Geiger 2001Go). In cultured mesangial cells, differences in expression of tensin were observed under different culture conditions. In the quiescent condition, expression of tensin might be stronger with changes in the structures. Although we did not examine the presence of FAs in the MCs, these findings suggest that MCs may contain FAs and control cell functions, including cell adhesion and signal transduction, in different ways from other glomerular cells under all conditions. In the future, it will be necessary to examine the presence of FAs and the expression of tensin under abnormal conditions in the MCs.

At present, two proteins are used as markers of MCs, i.e., {alpha}-SMA and Thy-1.1. {alpha}-SMA is detected in MCs during proliferative and regenerative processes after severe injury but is not detected in normal MCs (Johnson et al. 1991Go; Alpers et al. 1992Go; Utsunomiya et al. 1999Go; Kaneko et al. 2001Go). Thy-1.1 is expressed in rat MCs but not in other mammalian MCs (Ishizaki et al. 1980Go; Harada et al. 1982Go; Yamamoto and Wilson 1987Go; Holthofer et al. 1995Go). The specific expression of the protein in rat MCs in this study suggested that tensin can be used as a marker for rat MCs. We also identified the localization of tensin in human MCs by IHC using renal biopsy specimens from patients with chronic glomerulonephritis with or without MC proliferation (Yamashita et al. 2004Go). This finding suggests that tensin can also act as a marker protein for human MCs. In comparison with these two proteins, tensin has the advantage of detecting rat and human MCs under both normal and proliferative conditions.

In summary, it appears that tensin is related to MC attachment to the surrounding ECM, by which MCs, together with the GBM, form the basic structural stabilizing system of the glomerular capillary architecture. It was also suggested that the signal transduction regulated by tensin may be related to specific mechanisms of MC and matrix regeneration. Furthermore, tensin can act as a marker for rat and human MCs because the expression of tensin was detected only in MCs.


    Acknowledgments
 
We thank M. Yoshida, K. Sato, J. Nakamoto, T. Shibata, and T. Shigihara (Juntendo University; Tokyo, Japan) for their technical assistance.


    Footnotes
 
Received for publication September 3, 2003; accepted January 14, 2004


    Literature Cited
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 Summary
 Introduction
 Materials and Methods
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 Discussion
 Literature Cited
 

Alpers CE, Hudkins KL, Gown AM, Johnson RJ (1992) Enhanced expression of ‘muscle-specific’ actin in glomerulonephritis. Kidney Int 41:1134–1142[Medline]

Barners JL, Hevey KA, Hastings RR, Bocanegra RA (1994a) Mesangial cell migration precedes proliferation in Habu snake venom-induced glomerular injury. Lab Invest 70:460–467[Medline]

Barners JL, Hastings RR, De la Garza MA (1994b) Sequential expression of cellular fibronectin by platelets, macrophages, and mesangial cells in proliferative glomerulonephritis. Am J Pathol 145:585–597[Abstract]

Barners JL, Torres ES, Michell RJ, Peters JH (1995) Expression of alternatively spliced fibronectin variants during remodeling in proliferative glomerulonephritis. Am J Pathol 147:1361–1371[Abstract]

Bieritz B, Spessotto P, Colombatti A, Jahn A, Prols F, Hartner A (2003) Role of {alpha}8 integrin in mesangial cell adhesion, migration, and proliferation. Kidney Int 64:119–127[Medline]

Bradfield JW, Cattell V, Smith J (1977) The mesangial cell in glomerulonephritis. II. Mesangial proliferation caused by Habu snake venom in the rat. Lab Invest 36:487–492[Medline]

Busiek DF, Baragi V, Nehring LC, Parks WC, Welgus HG (1995) Matrilysin expression by human mononuclear phagocytes and its regulation by cytokines and hormones. J Immunol 154:6484–6491[Abstract/Free Full Text]

Cattell V, Bradfield JW (1977) Focal mesangial proliferative glomerulonephritis in the rat caused by Habu snake venom. A morphologic study. Am J Pathol 87:511–524[Abstract]

Chen H, Duncan IC, Bozorgchami H, Lo SH (2002) Tensin1 and a previously undocumented family member, tensin2, positively regulate cell migration. Proc Natl Acad Sci USA 99:733–738.[Abstract/Free Full Text]

Chen H, Ishii A, Wong WK, Chen LB, Lo SH (2000) Molecular characterization of human tensin. Biochem J 351:403–411.[Medline]

Chen H, Lo SH (2003) Regulation of tensin-promoted cell migration by its focal adhesion binding and Src homology domain 2. Biochem J 370:1039–1045[Medline]

Chuang JZ, Lin DC, Lin S (1995) Molecular cloning, expression, and mapping of the high affinity actin-capping domain of chicken cardiac tensin. J Cell Biol 128:1095–1109[Abstract]

Cortes–Hernandez J, Fossati–Jimach L, Carugati A, Potter PK, Walport MJ, Cook HT, Botto M (2002) Murine glomerular mesangial cell uptake of apoptotic cells is inefficient and involves serum-mediated but complement-independent mechanisms. Clin Exp Immunol 130:459–466[Medline]

Davis S, Lu ML, Lo SH, Lin S, Butter JA, Druker BJ, Roberts TM, et al. (1991) Presence of an SH2 domain in the actin-binding protein tensin. Science 252:712–715[Medline]

Diaz–Cazorla M, Perez–Sala D, Ros J, Jimenez W, Fresno M, Lamas S (1999) Regulation of cyclooxygenase-2 expression in human mesangial cells—transcriptional inhibition by IL-13. Eur J Biochem 260:268–274[Abstract/Free Full Text]

Drenckhahn D, Franke RP (1988) Ultrastructural organization of contractile and cytoskeletal proteins in glomerular podocytes of chicken, rat, and man. Lab Invest 59:673–682[Medline]

Drenckhahn D, Schnittler H, Nobiling R, Kriz W (1990) Ultrastructural organization of contractile proteins in rat glomerular mesangial cells. Am J Pathol 137:1343–1351[Abstract]

Dugina V, Fontao L, Chaponnier C, Vasiliev J, Gabbiani G (2001) Focal adhesion features during myofibroblastic differentiation are controlled by intracellular and extracellular factors. J Cell Sci 114:3285–3296[Medline]

Gauer S, Yao J, Schocklmann HO, Sterzel RB (1997) Adhesion molecules in the glomerular mesangium. Kidney Int 51:1447–1453[Medline]

Harada K, Yamamoto T, Hara M, Kihara I (1982) Antigenic association between kidney and thymocyte. Acta Pathol Jap 32:483–489[Medline]

Hartner A, Schocklmann H, Prols F, Muller U, Sterzel RB (1999) {alpha}8 integrin in glomerular mesangial cells and in experimental glomerulonephritis. Kidney Int 56:1468–1480[Medline]

Hiura K, Lim SS, Little SP, Lin S, Sato M (1995) Differentiation dependent expression of tensin and cortactin in chicken osteoclasts. Cell Motil Cytoskel 30:272–284[Medline]

Holthofer H, Sainio K, Miettinen A (1995) The glomerular mesangium: studies of its developmental origin and markers in vivo and in vitro. APMIS 103:354–366[Medline]

Hughes J, Lin Y, Van Damme J, Savill J (1997) Human glomerular mesangial cell phagocytosis of apoptotic neutrophils: mediation by a novel CD36-independent vitronectin receptor/thrombospondin recognition mechanism that is uncoupled from chemokine secretion. J Immunol 158:4389–4397[Abstract]

Ishii A, Lo SH (2001) A role of tensin in skeletal-muscle regeneration. Biochem J 356:737–745[Medline]

Ishizaki M, Sato S, Sano J, Fukuda Y, Sugisaki Y, Masugi Y (1980) The presence of Thy-1.1 antigen in rat glomerular mesangial cells. Biomed Res 1:438–442

Johnson RJ, Iida H, Alpers CE, Majesky MW, Schwartz SM, Pritzl P, Gordon K, Gown AM (1991) Expression of smooth muscle cell phenotype by rat mesangial cells in immune complex nephritis. {alpha}-Smooth muscle actin is a marker of mesangial cell proliferation. J Clin Invest 87:847–858[Medline]

Kaneko Y, Nakazawa K, Higuchi M, Hora K, Shigematsu H (2001) Glomerular expression of {alpha}-smooth muscle actin reflects disease activity of IgA nephropathy. Pathol Int 51:833–844[Medline]

Katz BZ, Zamir E, Bershadsky A, Kam Z, Yamada KM, Geiger B (2000) Physical state of the extracellular matrix regulates the structure and molecular composition of cell-matrix adhesions. Mol Biol Cell 11:1047–1060[Abstract/Free Full Text]

Kikkawa Y, Virtanen I, Miner JH (2003) Mesangial cells organize the glomerular capillaries by adhering to the G domain of laminin {alpha}5 in the glomerular basement membrane. J Cell Biol 161:187–196[Abstract/Free Full Text]

Kook S, Kim do H, Shim SR, Kim W, Chun JS, Song WK (2003) Caspase-dependent cleavage of tensin induces disruption of actin cytoskeleton during apoptosis. Biochem Biophys Res Commun 303:37–45[Medline]

Kriz W, Elger M, Lemley KV, Sakai T (1990a) Structure of the glomerular mesangium: a biomechanical interpretation. Kidney Int 38(suppl 30):S2–9

Kriz W, Elger M, Lemley KV, Sakai T (1990b) Mesangial cell-glomerular basement membrane connections counteract glomerular capillary and mesangium expansion. Am J Nephrol 10(suppl 1):4–13[Medline]

Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685[Medline]

Lemley KV, Elger M, Koeppen–Hagemann I, Kretzler M, Nagata M, Sakai T, Uiker S, et al. (1992) The glomerular mesangium: capillary support function and its failure under experimental conditions. Clin Invest 70:843–856[Medline]

Lo SH, An Q, Bao S, Wong WK, Lin Y, Janmey PA, Hartwig JH, et al. (1994a) Molecular cloning of chick cardiac muscle tensin. Full-length cDNA sequence, expression, and characterization. J Biol Chem 269:22310–22319[Abstract/Free Full Text]

Lo SH, Janmey PA, Hartwig JH, Chen LB (1994b) Interactions of tensin with actin and identification of its three distinct actin-binding domains. J Cell Biol 125:1067–1075[Abstract]

Lo SH, Weisberg E, Chen LB (1994c) Tensin: a potential link between the cytoskeleton and signal transduction. Bioessays 16: 817–823[Medline]

Lo SH, Yu QC, Degenstein L, Chen LB, Fuchs E (1997) Progressive kidney degeneration in mice lacking tensin. J Cell Biol 136:1349–1361[Abstract/Free Full Text]

Prols F, Hartner A, Schocklmann HO, Muller U, Sterzel RB (1999) Mesangial cells and their adhesive properties. Exp Nephrol 7:137–146[Medline]

Rovin BH, Lu L, Cosio A (2001) Cyclopentenone prostaglandins inhibit cytokine-induced nf-kappab activation and chemokine production by human mesangial cells. J Am Soc Nephrol 12:1659–1667[Abstract/Free Full Text]

Sakai T, Kriz W (1987) The structural relationship between mesangial cells and basement membrane of the renal glomerulus. Anat Embryol (Berl) 176:373–386[Medline]

Salgia R, Brunkhorst B, Pisick E, Li JL, Lo SH, Chen LB, Griffin JD (1995) Increased tyrosine phosphorylation of focal adhesion proteins in myeloid cell lines expressing p210BCR/ABL. Oncogene 11:1149–1155[Medline]

Schlondorff D, Mori T (1990) Contributions of mesangial cells to glomerular immume functions. Klin Wochenschr 68:1138–1144[Medline]

Striker GE, Lange MA, Mackay K, Bernstein K, Striker LJ (1987) Glomerular cells in vitro. Adv Nephrol Necker Hosp 16:169–186[Medline]

Towbin H, Staehelin T, Gordon J (1979) Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci USA 76:4350–4354[Abstract]

Utsunomiya Y, Kawamura T, Abe A, Imai H, Hirano K, Hosoya T, Sakai O (1999) Significance of mesangial expression of {alpha}-smooth muscle actin in the progression of IgA nephropathy. Am J Kidney Dis 34:902–910[Medline]

Yamamoto T, Wilson CB (1987) Complement dependence of antibody-induced mesangial cell injury in the rat. J Immunol 138:3758–3765[Abstract/Free Full Text]

Yamashita M, Horikoshi S, Asanuma K, Takahara H, Shirato I, Tomino Y (2004) Tensin is potentially involved in extracellular matrix production in mesangial cells. Histochem Cell Biol 121:245–254[Medline]

Zamir E, Geiger B (2001) Molecular complexity and dynamics of cell-matrix adhesions. J Cell Sci 114:3583–3590.[Medline]

Zamir E, Katz BZ, Aota S, Yamada KM, Geiger B, Kam Z (1999) Molecular diversity of cell-matrix adhesions. J Cell Sci 112: 1655–1669[Abstract/Free Full Text]

Zamir E, Katz M, Posen Y, Erez N, Yamada KM, Katz BZ, Lin S, Lin DC, et al. (2000) Dynamics and segregation of cell-matrix adhesions in cultured fibroblasts. Nature Cell Biol 2:191–196[Medline]





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