Alfa-smooth Muscle Actin Research Articles

Scopoletin alleviates high glucose-induced toxicity in human renal proximal tubular cells via inhibition of oxidative damage, epithelial-mesenchymal transition, and fibrogenesis.

Recent years of evidence suggest the crucial role of renal tubular cells in developing diabetic kidney disease. Scopoletin (SCOP) is a plant-based coumarin with numerous biological activities. This study aimed to determine the effect of SCOP on renal tubular cells in developing diabetic kidney disease and to elucidate mechanisms. In this study, SCOP was evaluated in vitro using renal proximal tubular (HK-2) cells under hyperglycemic conditions to understand its mechanism of action. In HK-2 cells, SCOP alleviated the high glucose-generated reactive oxygen species (ROS), restored the levels of reduced glutathione, and decreased lipid peroxidation. High glucose-induced alteration in the mitochondrial membrane potential was markedly restored in the SCOP-treated cells. Moreover, SCOP significantly reduced the high glucose-induced apoptotic cell population in the Annexin V-FITC flow cytometry study. Furthermore, high glucose markedly elevated the mRNA expression of fibrotic and extracellular matrix (ECM) components, namely, transforming growth factor (TGF)-β, alfa-smooth muscle actin (α-SMA), collagen I, and collagen III, in HK-2 cells compared to the untreated cells. SCOP treatment reduced these mRNA expressions compared to the high glucose-treated cells. Collagen I and TGF-β protein levels were also significantly reduced in the SCOP-treated cells. Further findings in HK-2 cells revealed that SCOP interfered with the epithelial-mesenchymal transition (EMT) in the high glucose-treated HK-2 cells by normalizing E-cadherin and downregulating the vimentin and α-SMA proteins. In conclusion, SCOP modulates the high glucose-generated renal tubular cell oxidative damage and accumulation of ECM components and may be a promising molecule against diabetic nephropathy.

α-Smooth Muscle Actin as Predictors of Early Recurrence in Early-Stage Ductal Type Breast Cancer After Mastectomy and Chemotherapy.

Breast cancer recurrence after surgery was a sign that the progress of the disease was continuing. Early detection of breast cancer patients who are at risk requires development of a marker. Alfa smooth muscle actin (α-SMA) plays a role in the local recurrence process of invasive ductal carcinoma (IDC). Currently, existing tumor markers are used to predict the prognosis of breast cancer in general, not the early stages. Therefore, it was thought that finding α-SMA expression might predict early recurrence in early-stage IDC more accurately than others. This study investigated the potential role of α-SMA expression as a predictor of early recurrence in early-stage IDC and its relationship to clinicopathological factors. The study design was cross-sectional, with data obtained from the medical records of Dr. Koesnadi, General Hospital, Bondowoso, Indonesia. Bivariate and multivariate analysis was performed to analyze data. We included 50 subjects divided into the local recurrence group (n=25) and the non-local recurrence group (n=25). We found a statistically significant correlation between the incidence of local recurrence in early-stage IDC and the high expression of α-SMA (odd ratio [OR]=23.22, 95% confidence interval [CI]=5.101-105.7, P=0.001). Clinicopathological variables and α-SMA expression did not have a significant correlation. In early-stage IDC, α-SMA expression had the potential to predict and could be an independent prognostic factor for early recurrence.

Effects of Dipeptidyl Peptidase-4 Inhibitor and Angiotensin-Converting Enzyme Inhibitor on Experimental Diabetic Kidney Disease

Diabetic kidney disease (DKD) is the leading cause of end-stage kidney disease in the United States and worldwide. Proteinuria is a major marker of the severity of injury. Dipeptidyl peptidase-4 inhibitor (DPP-4I) increases incretin-related insulin production and is, therefore, used to treat diabetes. We investigated whether DPP4I could have direct effect on kidney independent of its hypoglycemic activity. We, therefore, tested the effects of DPP4I with or without angiotensin-converting enzyme inhibitor (ACEI) on the progression of diabetic nephropathy and albuminuria in a murine model of DKD. eNOS−/−db/db mice were randomized to the following groups at age 10 weeks and treated until sacrifice: baseline (sacrificed at week 10), untreated control, ACEI, DPP4I, and combination of DPP4I and ACEI (Combo, sacrificed at week 18). Systemic parameters and urine albumin–creatinine ratio were assessed at baseline, weeks 14, and 18. Kidney morphology, glomerular filtration rate (GFR), WT-1, a marker for differentiated podocytes, podoplanin, a marker of foot process integrity, glomerular collagen IV, and alpha-smooth muscle actin were assessed at the end of the study. All mice had hyperglycemia and proteinuria at study entry at week 10. Untreated control mice had increased albuminuria, progression of glomerular injury, and reduced GFR at week 18 compared with baseline. DPP4I alone reduced blood glucose and kidney DPP-4 activity but failed to protect against kidney injury compared with untreated control. ACEI alone and combination groups showed significantly reduced albuminuria and glomerular injury, and maintained GFR and WT-1+ cells. Only the combination group had significantly less glomerular collagen IV deposition and more podoplanin preservation than the untreated control. DPP-4I alone does not decrease the progression of kidney injury in the eNOS−/−db/db mouse model, suggesting that targeting only hyperglycemia is not an optimal treatment strategy for DKD. Combined DPP-4I with ACEI added more benefit to reducing the glomerular matrix.

Biochanin A Ameliorates Nephropathy in High-Fat Diet/Streptozotocin-Induced Diabetic Rats: Effects on NF-kB/NLRP3 Axis, Pyroptosis, and Fibrosis.

Nephropathy is the most prevalent microvascular disorder in diabetes mellitus. Oxidative stress and inflammatory cascade provoked by the persistent hyperglycemic milieu play integral roles in the aggravation of renal injury and fibrosis. We explored the impact of biochanin A (BCA), an isoflavonoid, on the inflammatory response, nod-like receptor protein 3 (NLRP3) inflammasome activation, oxidative stress, and fibrosis in diabetic kidneys. A high-fat-diet/streptozotocin (HFD/STZ)-induced experimental model of diabetic nephropathy (DN) was established in Sprague Dawley rats, and in vitro studies were performed in high-glucose-induced renal tubular epithelial (NRK-52E) cells. Persistent hyperglycemia in diabetic rats was manifested by perturbation of renal function, marked histological alterations, and oxidative and inflammatory renal damage. Therapeutic intervention of BCA mitigated histological changes, improved renal function and antioxidant capacity, and suppressed phosphorylation of nuclear factor-kappa B (NF-κB) and nuclear factor-kappa B inhibitor alpha (IκBα) proteins. Our in vitro data reveal excessive superoxide generation, apoptosis, and altered mitochondrial membrane potential in NRK-52E cells that were cultured in a high-glucose (HG) environment were subsided by BCA intervention. Meanwhile, the upregulated expressions of NLRP3 and its associated proteins, the pyroptosis-indicative protein gasdermin-D (GSDMD) in the kidneys, and HG-stimulated NRK-52E cells were significantly ameliorated by BCA treatment. Additionally, BCA blunted transforming growth factor (TGF)-β/Smad signaling and production of collagen I, collagen III, fibronectin, and alfa-smooth muscle actin (α-SMA) in diabetic kidneys. Our results indicate the plausible role of BCA in attenuating DN, presumably through modulation of the apoptotic cascade in renal tubular epithelial cells and the NF-κB/NLRP3 axis.

ADAMTS13 Plays a Critical Role in Acetaminophen-Induced Acute Liver Injury in Mice, and Can be a Novel Therapeutic Option

【Background】 Acetaminophen (APAP) is one of the major causes of drug-induced acute liver injury. It is known that VWF deficiency reduces hepatic platelet accumulation and promotes liver repair in APAP-induced acute liver injury in mice. We hypothesized that VWF-ADAMTS13 axis plays a role in the pathophysiology of APAP-induced acute liver injury. To investigate this hypothesis, we have used a mouse experimental model of APAP-induced hepatic injury. 【Method】 The extent of APAP-induced liver injury was compared between wild-type (WT) and ADAMTS13 gene-deleted (KO) mice (C57BL/6 background). At first, mice were fasted for 24 hours before administration of 300 mg/kg APAP via intraperitoneal injection. Forty-eight hours after APAP administration, Hepatic blood flow was measured as the average of the surface blood flow on the anterior and inferior surfaces of the liver in the right and left lobes using Laser Doppler flowmetry (ALF21N, Advance Co, Tokyo, Japan), followed by sampling of blood and hepatic tissue. Blood cell counts and biochemical analysis including aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were performed. Also, histological analysis and gene expression analysis by quantitative PCR (qPCR) such as interleukin-6 (IL-6), interleukin-1beta (IL1-β), tumor necrosis factor-alfa (TNF-α), Keratinocyte-derived chemokines (KC), monocyte chemoattractant protein-1 (MCP-1), transforming growth factor-beta (TGF-β), alfa-smooth muscle actin (α-SMA) and collagen type I alfa-1 (COL1α1) were conducted using the liver samples. In addition, to confirm whether recombinant ADAMTS13 has a protective effect against APAP-induced liver injury, we administered recombinant ADAMTS13 to WT mice through the retro-orbital cavity 30 min before APAP administration and examined whether the liver injury was reduced. 【Result】 In APAP-challenged KO mice (N=13), serum levels of AST and ALT were significantly elevated compared to WT mice (N=9)（AST; 3115.4 vs 1331.1 IU/L, ALT; 3911.5 vs 1647.8 IU/L）. The area of liver necrosis was increased in KO mice compared to WT mice （area of hepatic necrosis; 58.0 vs 42.9 %）. In addition, inflammation markers other than TNF-α and MCP-1 were not significantly different, but TNF-α and MCP-1 were significantly elevated in KO mice compared to WT mice. Furthermore, hepatic blood flow was decreased in KO mice compared to WT mice（18.3 vs 21.6 ml/hr）, in accordance with the significantly reduced levels of the numbers of platelets in KO mice（269.3 vs 503.2 x109/L）. Surprisingly, the increases of ALT values and the area of hepatic necrosis observed in WT mice were significantly suppressed by recombinant human ADAMTS13 administration. 【Discussion】 We have shown that APAP-induced liver injury is exacerbated by enhanced inflammation in KO mice and clarified that ADAMTS13 plays a critical role in the liver injury induced by APAP. Moreover, we found that recombinant human ADAMTS13 administration reduces APAP-induced hepatic injury. Our results suggest that recombinant human ADAMTS13 can be a novel therapeutic option for APAP poisoning.

Culturing of Cardiac Fibroblasts in Engineered Heart Matrix Reduces Myofibroblast Differentiation but Maintains Their Response to Cyclic Stretch and Transforming Growth Factor β1.

Isolation and culturing of cardiac fibroblasts (CF) induces rapid differentiation toward a myofibroblast phenotype, which is partly mediated by the high substrate stiffness of the culture plates. In the present study, a 3D model of Engineered Heart Matrix (EHM) of physiological stiffness (Youngs modulus ~15 kPa) was developed using primary adult rat CF and a natural hydrogel collagen type 1 matrix. CF were equally distributed, viable and quiescent for at least 13 days in EHM and the baseline gene expression of myofibroblast-markers alfa-smooth muscle actin (Acta2), and connective tissue growth factor (Ctgf) was significantly lower, compared to CF cultured in 2D monolayers. CF baseline gene expression of transforming growth factor-beta1 (Tgfβ1) and brain natriuretic peptide (Nppb) was higher in EHM-fibers compared to the monolayers. EHM stimulation by 10% cyclic stretch (1 Hz) increased the gene expression of Nppb (3.0-fold), Ctgf (2.1-fold) and Tgfβ1 (2.3-fold) after 24 h. Stimulation of EHM with TGFβ1 (1 ng/mL, 24 h) induced Tgfβ1 (1.6-fold) and Ctgf (1.6-fold). In conclusion, culturing CF in EHM of physiological stiffness reduced myofibroblast marker gene expression, while the CF response to stretch or TGFβ1 was maintained, indicating that our novel EHM structure provides a good physiological model to study CF function and myofibroblast differentiation.

TUMOR NECROSIS FACTOR (TNF)-α PLAYS A ROLE IN ORAL ULCERATION IN RATS

<h3>Objectives</h3> Evaluate the role of tumor necrosis factor (TNF)-α in the healing process of oral traumatic ulcers (OTUs) in rats. <h3>Study Design</h3> The animals were randomly divided into 6 experimental groups: 1 treated intravascularly with saline solution and 5 treated with infliximab at Infliximab (INF)-1, INF-INF-3, INF-5, INF-7, or INF-10 mg/kg. The animals were euthanized on days 1, 3, 7, 14, and 21 after ulceration. The ulcers were clinically measured (wound area) and the mucosa samples were histologically (scores 0-4), histomorphometrically (cell counting polymorphonuclear cells), and immunohistochemically (scores 0-3 for TNF-α and alfa-smooth muscle actin (α-SMA)) analyzed. Evans blue assay was performed to measure vascular permeability. One- and two-way analyses of variance and Kruskal-Wallis tests were performed (GraphPad Prism 5.0, <i>P</i> < .05). <h3>Results</h3> There was a reduction in the OTU area of the infliximab-treated groups (INF-5, INF-7, and INF-10) on day 1 (<i>P</i> = .043) and Evan blue extravasation (<i>P</i> = .032), polymorphonuclear cell counts (<i>P</i> = .001), and TNF-α immunostaining (<i>P</i> = .025) were significantly reduced in the dose INF-5 group compared to the saline group on the same day. In day 3, all INF-treated groups showed lower scores than the saline group (<i>P</i> = .023), but α-SMA immunostaining was reduced in day 7 in the INF-treated groups (<i>P</i> = .006). <h3>Conclusions</h3> TNF-α blockade reduced acute inflammatory process and delay wound healing in OTU.

Effects of topical docosahexaenoic acid on postoperative fibrosis in an animal model of glaucoma filtration surgery.

The aim of this study was to evaluate docosahexaenoic acid (DHA) as a potential antifibrotic agent after glaucoma filtration surgery (GFS) in rats. A total of 36 10-week-old Brown Norway rats underwent GFS. Animals were equally divided into three groups: a control group, a DHA group and a mitomycin C (MMC) group. Intraocular pressure (IOP) was measured using a dynamic rebound tonometer, and a photograph of the surgical site was taken on days 1, 3, 7, 10, 14 and 17. The incorporation of DHA into fibroblasts was evaluated by gas chromatography. The expression of alfa-smooth muscle actin (α-SMA) and Smad proteins was assessed by Western blotting. IOP decreased after surgery in animals from the three groups on day 1 after surgery. Over time, IOP remained lower in the DHA and MMC groups than in the control group (median [interquartile range] 8.0 [7.0-8.0] and 8.0 [7.3-8.0] mmHg vs. 9.0 [8.0-9.0] mmHg, respectively; p < 0.001). Bleb area in the DHA and MMC groups remained larger than that of the control group from day 7 to day 14 (3.9 [2.9-5.2] and 3.5 [2.3-4.4] mm2 vs. 2.3 [2.0-2.8] mm2 , respectively; p=0.0021). We did not observe any change in DHA concentrations in the fibroblasts of the DHA group compared with the other groups. The impact of DHA on IOP and bleb area was similar to that of MMC. The mechanisms of action of DHA in rat eye fibroblasts deserve further investigation.

Decellularized bovine aorta as a promising 3D elastin scaffold for vascular tissue engineering applications.

Aim: To evaluate the suitability of using aorta elastin scaffold, in combination with human adipose-derived mesenchymal stem cells (hAd-MSCs), as an approach for cardiovascular tissue engineering. Materials & Methods: Human adipose-derived MSCs were seeded on elastin samples of decellularized bovine aorta. The samples were cultured in vitro to investigate the inductive effects of this scaffold on the cells. The results were evaluated using histological, and immunohistochemical methods, as well as MTT assay, DNA content, reverse transcription-PCR and scanning electron microscopy. Results: Histological staining and DNA content confirmed the efficacy of decellularization procedure (82% DNA removal). MTT assay showed the construct's ability to support cell viability and proliferation. Cell differentiation was confirmed by reverse transcription-PCR and positive immunohistochemistry for alfa smooth muscle actin and von Willebrand. Conclusion: The prepared aortic elastin samples act as a potential scaffold, in combination with MSCs, for applications in cardiovascular tissue engineering. Further experiments in animal models are required to confirm this.

Generation of transgene-free induced pluripotent stem cells from cardiac fibroblasts of goat embryos.

Induced pluripotent stem cells (iPSCs) hold a great potential for therapeutic regenerative medicine. The aim of this study was to generate induced pluripotent stem cells from goat embryonic cardiac tissue derived fibroblasts. The isolated cardiac fibroblasts from the cardiac tissue of goat embryos were positive for alfa smooth muscle actin, vimentin and discoidin domain receptor2. From these cells, we generated transgene free iPSCs using piggyBac transposons / transposase using five transcription factors (Oct4, Sox2, Klf, Myc and Lin 28). The generated iPSCs were SSEA1, SSEA4 and Oct4 positive. They were cultured on neofeeders using 20% Serum replacement - IMDM with bFGF. They could form cystic and compact embryoid bodies that showed differentiated ectodermal and mesodermal like cells when cultured using 20% FBS-IMDM without bFGF. The iPSCs, generated in the frame of this approach were produced without the use of integrating virus and the reprogramming transgenes were removed at the end of the process. Though there were limitations in the approach used, a substantial sign of reprogramming was obtained.

LEIOMYOSARCOMA OF THE OVARIAN VEIN WITH THE RENAL VEIN INVASION: A CASE REPORT

A 56-year-old woman had complained of two year's consecutive left-side abdominal pain. Retroperitoneal tumor was diagnosed and the patient was referred to our institute in December 2014. Laboratory data including endocrinological activity and serological markers were within the normal ranges. Imaging studies showed that the solid tumor measuring 5 cm in diameter was uncovered in the retroperitoneum, between the abdominal aorta and left kidney. The patient underwent surgical removal of the tumor with the left kidney because the mass was highly adhesive to the left ovarian vessels and left renal vein. Histological examination showed proliferating spindle cells in the tumor, and immunoreactivity for desmin and alfa-smooth muscle actin in tumor cells confirmed the diagnosis of leiomyosarcoma originated in left ovarian vein with left renal vein invasion. The patient has been free of disease 21 months after the surgery. Ovarian vein leiomyosarcoma is extremely rare and we have found 18 cases in literature. Furthermore, only three cases of leiomyosarcoma arising from the ovarian vein with the renal vein invasion were reported including our case.

Extracellular matrix remodeling in equine sarcoid: an immunohistochemical and molecular study.

BackgroundEquine sarcoids are locally invasive, fibroblastic benign skin tumors. Bovine papillomavirus type-1 (BPV-1) and/or Bovine papillomavirus type-2 (BPV-2) are believed to be the causative agent of sarcoids, although the mechanisms by which the virus induce the tumor are still poorly understood. We hypothesized that in genetically predisposed equines latent BPV infection may be reactivated by immunosoppression and/or mechanical injury leading to a form of pathologic wound which may transform into a sarcoid. In this study, we investigated in 25 equine sarcoids and in five normal skin samples the histological features and evaluated the immunohistochemical and molecular expression of type I and type III Collagen, vimentin (VIM), alfa Smooth Muscle Actin (α-SMA), Matrix Metalloproteinase (MMPs) -2, 9, 14 and tissue inhibitor of metalloproteinase 2 (TIMP-2).ResultsIn 64 % of investigated sarcoids, type I collagen staining was stronger than that of type III collagen. In 80 % of sarcoids, SFs were strongly positive for vimentin and negative for α-SMA; the remaining sarcoid samples (20 %) showed 70–80 % of SFs labeled for vim and approximately 20–30 % labeled for α-SMA. Moreover, all sarcoid specimen showed a variable staining pattern (weak to moderate) for MMP-9 and MMP-14, and a moderate to strong staining for MMP-2 and TIMP-2. Biochemical analysis confirmed immunohistochemical results and showed in sarcoids, for the first time, the cleaved form of MMP9, the 35 KDa active species for MMP-9.ConclusionsThis study revealed that in equine sarcoids exhibit an altered turnover of the Extracellular Matrix (ECM) deposition and degradation, as result of an altered expression of MMPs and TIMPs. Therefore, these observations seem to confirm that the basic mechanism for growth of equine sarcoids could be a neoplastic transformation during wound healing.

Congenital Granular Cell Lesion in the Ventral Tongue in a 2 Day-Old Newborn

A 2-day-old newborn healthy girl from Cancun, Quintana Roo, with a polypoid mass in the ventral tongue near the Blandin Nun salivary glands, this is the first case in the family with this pathology. The mass is peduculated, exophytic, smooth, soft and the same color of the mucosa, measuring 8 × 6 × 4 mm., and the clinical diagnosis was mucocele versus hamartomas or coristoma. The excisional biopsy was made under local anesthesia, not complications were present during the surgical removal. Microscopically stained with H&E the lesion was composed of large cell containing abundant granular cytoplasm and small hyperchromatic nuclei. The immmunohistochemical was positive for vimetine, but negative for S-100 protein, alfa-smooth muscle actin an CD68. The diagnosis was CONGENITAL GRANULAR CELL LESION (Histological classification by the WHO) because the origin is the soft tissues and not in the alveolar regions, there are only 10 cases reported in the literature, the first was diagnosed in 1975 Dixter CT.

Morphological evaluation of metastatic cholangiocarcinoma in a dog

Cholangiocellular carcinomas are aggressive, malignant neoplasms of biliary epithelium usually arising from intrahepatic bile ducts. The aim of this study was to describe the immunohistochemical characteristics of metastatic ductular/papillary cholangiocarcinoma in a common breed dog. A 16-year old common breed dog was submitted for necropsy with an anamnesis of apathy, vomiting, anorexia and intermittent neurological symptoms. After complete necropsy, samples were taken and processed using paraffin technique. The sections were stained with Hematoxylin-eosin, Masson’s trichrome, and PAS methods. Immunohistochemical examination was also performed using alpha smooth muscle actin, multicytokeratin AE1/AE3, cytokeratin 7, cytokeratin 19 and vimentin antibodies. At necropsy multiple masses in the liver parenchyma, and several metastases in the lungs, pancreas, intestinal serosa, omentum and on the surface of the diaphragm muscle were observed. Histologically the tumor consisted of pleomorphic epithelial cells, arranged in ductules, presenting intraluminal papillary growth pattern and marked fibroplasia. Vascular invasion, intra and extrahepatic metastases were also present. The tumoral cells were AE1/AE3, cytokeratin 7 and cytokeratin 19 positive, alfa smooth muscle actin and vimentin negative. Immunohistochemical exam proved to be useful in differentiating cholangiocellular carcinoma from hepatocellular carcinoma and several extrahepatic metastatic carcinomas.

Expression of gap junction proteins connexins 26, 30, and 43 in Dupuytren’s disease

Background and purpose Dupuytren’s disease (DD) is a benign fibroproliferative process of the palmar aponeurosis showing similarities to wound healing. Communication of cells involved in wound healing is mediated by the composition of gap junction (GJ) proteins. We investigated the expression of 3 GJ proteins, connexins 26, 30, and 43 (Cx26, Cx30, and Cx43) in DD.Patients and methods Fragments of Dupuytren’s tissue from 31 patients (mean age 56 (30–76) years, 24 male) were analyzed immunohistochemically and compared to control tissue for expression of the GJ proteins Cx26, Cx30, and Cx43 and also alfa-smooth muscle actin (α-SMA).Results 14 of 31 samples could be attributed to the involutional phase (α-SMA positive) whereas 17 samples had to be considered cords in the residual phase (α-SMA negative). Expression of Cx26 and Cx43 was seen in 12 of the 14 samples from the involutional phase, and Cx30 was seen in 7 of these. Only 4 of the 17 samples from the residual phase showed any Cx, and there was none in the controls.Interpretation The high expression of GJ proteins Cx26, Cx30, and Cx43 in α-SMA positive myofibroblast-rich nodules, which are characteristic of the active involutional phase of DD, suggests that connexins could be a novel treatment target for the treatment of DD.

Expression of VEGF, its receptors, and HIF-1α in Dupuytren’s disease

Background and purpose — Dupuytren’s disease (DD) is a benign fibroproliferative process that affects the palmar fascia. The pathology of DD shows similarities with wound healing and tumor growth; hypoxia and angiogenesis play important roles in both. We investigated the role of angiogenic proteins in DD.Patients and methods — The expression of vascular endothelial growth factor (VEGF), its receptors vascular endothelial growth factor receptor 1 (VEGFR1) and vascular endothelial growth factor receptor 2 (VEGFR2), hypoxia-inducible factor alfa (HIF-1α), and alfa-smooth muscle actin (α-SMA) were analyzed immunohistochemically in fragments of excised Dupuytren’s tissue from 32 patients. We compared these values to values for expression in a control group.Results — 15 of 32 samples could be attributed to the involutional phase (α-SMA positive), whereas 17 samples were considered to be cords at the residual phase (α-SMA negative). In the involutional phase, the HIF-1α and VEGFR2 expression was statistically significantly higher than in the residual phase and in the controls.Interpretation — Both the VEGFR2 receptor and HIF-1α were expressed in α-SMA positive myofibroblast-rich nodules with characteristics of DD in the active involutional phase. Thus, hypoxia and (subsequently) angiogenesis may have a role in the pathophysiology of DD.

The herbal medicine Inchinko-to reduces hepatic fibrosis in cholestatic rats

Several studies have reported the herbal medicine Inchinko-to (ICKT) to have an antifibrotic effect which thus leads to an improvement of hepatic injury. However, there are still few reports of its use in the treatment of cholestatic liver disorder. The aim of this study was to clarify whether the administration of ICKT will ameliorate hepatic fibrosis and injury in cholestatic rats. We performed bile duct ligation on 7-week-old male cholestatic Wistar rats and assigned them to one of three groups according to the method of treatment: (1) the SHAM group, (2) the NT-group (non-treatment group), and (3) the T-group (treatment-group). Rats in the T-group were orally administered ICKT (TJ-135) at a dose of 1 g/kg/day and were killed on the 17th postoperative day. We subsequently investigated the levels of fibrosis and various clinical markers through measurement of the following: serum levels of AST and ALT; tissue transforming growth factor-beta 1 (TGF-beta1); tissue inhibitor metalloprotease-1 mRNA (TIMP-1 mRNA) through real-time PCR analysis; and Azan staining and immunohistochemical staining of alfa-smooth muscle actin (alfa-SMA) to evaluate the degree of fibrosis. The levels of serum AST, serum ALT, and TGF-bata1 in the T-Group were significantly lower than those in the NT-Group. In addition, staining of Azan and alfa-SMA in the T-Group was significantly lower than those in the NT-Group. ICKT may help reduce hepatic fibrosis and injury by controlling stellate cell activation.

Primary Cutaneous PEComa

A 48-year-old woman attended a physician because of a solitary cutaneous nodule on the left lower leg. Microscopic examination of the excisional specimen revealed a dermal tumor composed of nests of epithelioid cells exhibiting clear cytoplasm. They had centrally located vesicular nuclei with distinct nucleoli. A rich network of capillaries was present throughout. The tumor showed an infiltrative border. There was no epidermal involvement. Periodic acid-Shif (PAS) and PAS-Diastase stains demonstrated glycogen deposition within the cytoplasm of the clear cells. Immunohistochemical evaluation revealed that the tumor cells were positive for HMB-45 and microftalmia associated transcription factor (MITF). Focal desmin positivity was also seen. The tumor cells were negative for S-100 protein, alfa smooth muscle actin, HHF-35, and various cytokeratins. The case is one of a primary cutaneous pecoma. Pecomas are rare, recently described mesenchymal tumors composed of perivascular epithelioid cells. They constitute a spectrum of lesions in different organs including angiomyolipoma of the kidney and liver, sugar tumor of the lung, lymphangiomatosis, and lymphangiomyoma. Primary cutaneous PEComas are exceptionally rare and have only recently been recognized. To date, these are approximately 22 cases in the English literature. Follow-up data is limited but they appear to behave in a benign fashion. We report an additional case with the goal of alerting dermatopathologists to this distinctive unusual neoplasm.

Alterations in proteoglycan components and histopathology of the peritoneum in uraemic and peritoneal dialysis (PD) patients

Proteoglycans (PGs) are dominant extracellular matrices (ECMs) in the peritoneal tissues. Human peritoneal mesothelial cells synthesize small proteoglycans including decorin. Peritonitis and long-term peritoneal dialysis (PD) cause fibrotic changes in the peritoneum that result in ECM remodelling and PG synthesis. Twenty-five peritoneal tissues from eight patients at initiation of PD, five long-term PD (>6 years) patients with severe peritonitis lasting for almost 1 month, nine patients after long-term PD (>6 years) without peritonitis and three normal subjects were included in the present study. Expressions of decorin, versican, hyaluronan, MMP-2, alfa smooth muscle actin (alphaSMA) and CD68 for macrophages in these specimens were examined by immunohistochemical staining. Although expression of decorin was detected in normal subjects, it was markedly decreased with long-term PD treatment. In long-term PD patients, the expression of versican was observed in their fibrotic-thickened peritoneum. Versican was present in fibrous regions, elastic lamina of the peritoneum, vascular walls and perivascular regions. Hyaluronan was observed in the whole thickened peritoneum, but its distribution differed in part from that of versican. MMP-2 was mainly observed around the blood vessels. Alfa SMA-positive cells, namely 'myofibroblasts' and CD68-positive cells, i.e. macrophages, were observed in the fibrotic-thickened peritoneum of long-term PD patients. Expressions of MMP-2, hyaluronan, SMA and CD68 in the peritoneum were marked in long-term PD patients' samples, which were strongly immunostained by versican, and were especially high in peritonitis patients. It appears that alterations in PGs, including marked induction of versican with peritonitis and disappearance of decorin, are involved in peritoneal remodelling in PD patients. Versican expression was closely related to the appearance of myofibroblasts and macrophages. These observations suggest that the alteration in PG components following PD therapy and severe inflammation contribute to fibrous thickening of the peritoneum.

Changes in expression of fibrotic markers and histopathological alterations in kidneys of mice chronically exposed to low and high Cd doses

The main target organ for cadmium (Cd) is the kidney, and more specifically the proximal tubular cells. Little is known about the effects of a long-term Cd exposure on the ultrastructure of the kidney and the involvement in tubulointerstitial fibrosis. Therefore, mice were exposed to Cd concentrations varying from 10 to 500 mg CdCl 2/l in the drinking water during 4, 16 and 23 weeks. Ultrastructural changes were studied by means of light- and electron microscopical analyses. Furthermore, the expression of the extracellular matrix (ECM) proteins collagen I and fibronectin, and the myofibroblast/epithelial-to-mesenchymal transition (EMT) marker alfa-smooth muscle actin (α-SMA) were studied by means of immunohistochemistry. The histopathological changes caused by Cd varied considerably from one animal to another, and from one individual cell to another. An exposure to Cd concentrations up to 100 mg CdCl 2/l elicited only minor changes that were restricted to increasing amounts of lysosomes and vacuolisation. When higher Cd concentrations were applied, the changes became more pronounced and featured mitochondrial damage, cellular swelling and loss of basal invaginations. An overproduction of the interstitial matrix component fibronectin and the expression of the myofibroblasts/EMT marker α-SMA in kidneys of mice exposed to 100 mg CdCl 2/l clearly indicated that an exposure to relatively low Cd doses might lead ultimately to renal fibrosis. Increasing the Cd dose (up to 500 mg CdCl 2/l) evoked an increased immunoreactivity for fibrotic markers. In conclusion we may state that concentrations up to 100 mg CdCl 2/l evoked minor changes, although the expression of fibrotic markers was increased. Changes became more pronounced when exposing to higher Cd concentrations.