Our study examined whether human bone marrow-derived MSCs are able to differentiate, in vitro, into functional epithelial-like cells. MSCs were isolated from the sternum of 8 patients with different hematological disorders. The surface phenotype of these cells was characterized.To induce epithelial differentiation, MSCs were cultured using Epidermal Growth Factor, Keratinocyte Growth Factor, Hepatocyte Growth Factor and Insulin-like growth Factor-II. Differentiated cells were further characterized both morphologically and functionally by their capacity to express markers with specificity for epithelial lineage. The expression of cytokeratin 19 was assessed by immunocytochemistry, and cytokeratin 18 was evaluated by quantitative RT-PCR (Taq-man). The data demonstrate that human MSCs isolated from human bone marrow can differentiate into epithelial-like cells and may thus serve as a cell source for tissue engineering and cell therapy of epithelial tissue.

Ochratoxin A (OTA) is considered to be a possible human urinary tract carcinogen, based largely on a rat model, but no molecular genetic changes in the rat carcinomas have yet been defined. The phosphorylated-S6 ribosomal protein is a marker indicating activity of the mammalian target of rapamycin, which is a serine/threonine kinase with a key role in protein biosynthesis, cell proliferation, transcription, cellular metabolism and apoptosis, while being functionally deregulated in cancer. To assess p-S6 expression we performed immunohistochemistry on formalin-fixed and paraffin-embedded tumours and normal tissues. Marked intensity of p-S6 expression was observed in highly proliferative regions of rat renal carcinomas and a rare angiosarcoma, all of which were attributed to prolonged exposure to dietary OTA. Only very small OTA-generated renal adenomas were negative for p-S6. Examples of rat subcutaneous fibrosarcoma and testicular seminoma, as well as of normal renal tissue, showed no or very weak positive staining. In contrast to the animal model, human renal cell carcinoma, upper urinary tract transitional cell carcinoma from cases of Balkan endemic nephropathy, and a human angiosarcoma were negative for p-S6. The combined findings are reminiscent of constitutive changes in the rat tuberous sclerosis gene complex in the Eker strain correlated with renal neoplasms, Therefore rat renal carcinogenesis caused by OTA does not obviously mimic human urinary tract tumourigenesis.

CD34, traditionally a marker of hematopoietic stem cells (HSCs), was found on endothelial cells and fibroblasts as well. At the level of the extraglomerular or intraglomerular mesangium, CD34 may signal either the presence of HSCs or, conversely, may be a marker of transdifferentiation. CD34-positive cells of the extraglomerular mesangium could migrate into the intraglomerular mesangium and participate in reparative processes at this level. The aim of our study was to analyze the presence of CD34 at the level of the extraglomerular and intraglomerular mesangium and its relationship with histological markers of activity and chronicity, as well as with other immunohistochemical markers in glomerulonephritis (GN). A cross-sectional study of 36 patients with GN was conducted. Conventional stains: hematoxylin-eosin, periodic acid Schiff, and Trichrome Gömöri, as well as immunohistochemistry: CD34, alpha smooth muscle actin (alpha SMA), vimentin, and proliferating cell nuclear antigen (PCNA) were employed. Activity and chronicity of GN were evaluated according to a scoring system initially used for lupus nephritis and antineutrophil-cytoplasmic-antibody-associated vasculitis. Immunohistochemistry was assessed using a semiquantitative score. The mean age was 46.44 +/- 12.97 years; 22 were male and 14 were female. The extraglomerular mesangium was visible on specimens in 30 patients. CD34 was present in the extraglomerular mesangium in 15 patients: 11 of these patients showed concomitant intraglomerular and extraglomerular mesangial CD34 immunostaining, while four showed only extraglomerular mesangial immunostaining. In three patients, CD34 immunostaining was present only in the intraglomerular mesangium. Twelve patients showed negative immunostaining in both the extraglomerular and the intraglomerular mesangium. Overall, there was a fair degree of relationship, which did not reach statistical significance between CD34 in the extraglomerular mesangium and CD34 in the intraglomerular mesangium across the 36 patients. In the intraglomerular mesangium, CD34 did not significantly correlate with mesangial alpha SMA, vimentin, PCNA, and activity or chronicity index. In the extraglomerular mesangium, CD34 did not show a significant correlation with alpha SMA, vimentin, or PCNA. The activity index and the chronicity index showed a good correlation with serum creatinine. Mesangial cell proliferation correlated well with the mesangial matrix increase, while interstitial vimentin showed a good correlation with interstitial alpha SMA. We demonstrated the presence of CD34 in the extraglomerular mesangium, which could be related to transdifferentiated mesangial cells or to HSCs in the absence of blood vessels at this level. Our study shows the value of histological indices for evaluating GN but cannot assign significance to CD34 immunolabeling for the assessment of GN.

DNA ploidy measurement has been applied uniquely to wax-embedded tissue of primary renal cell and metastatic tumours of a key experimental researcher on porcine ochratoxicosis, a control, and four transitional cell carcinomas from cases of Balkan endemic nephropathy. Primary renal tumour was diploid, and hyperdiploid metastasis was within the lower ploidy range for typical renal cell carcinoma. Three Balkan primary tumours showed extensive aneuploidy indicating marked nuclear instability, similar to model rat renal carcinoma caused by ochratoxin A. In contrast, much less nuclear instability in the putative occupational ochratoxicosis case fitted poorly with the ochratoxin A model.

Experimental renal cancer caused by ochratoxin A (OTA) in rats was first defined in the US National Toxicology Program (1989) and raised questions about any aetiological role in human urinary tract tumours. A review of histopathology in several rat kidney tumours from dietary OTA in recently described London studies, augmented by clinical immunohistochemistry for the first time for this mycotoxin, establishes their renal tubular cell origin. It had been assumed that the toxin might cause the human urothelial tumours associated with Balkan endemic nephropathy, but the present study could not support this. Comparison with a similar review of a metastasising renal tumour from a female rat of the NTP study consistently shows the kidney as the primary carcinogenic site for OTA. Morphological heterogeneity of these kidney tumours as epithelioid and/or sarcomatoid is revealed. Leiomyosarcoma was also diagnosed, and rhabdomyosarcoma differentiation was observed in the exceptionally aggressive NTP female tumour. The present pilot study involving immunohistochemistry indicates need for wider review of archived tumours for experimental evidence before formulating any epidemiological basis from a rat model for OTA’s relevance to idiopathic human renal cell carcinoma. Although the NTP study concluded that females are less sensitive to OTA than males, some female tumours still had heterogeneous morphology.

Endothelial cells (ECs) are active participants of an inflammatory process in glomeruli. EC damage has been shown to play an important role in the progression of glomerulonephritis (GN). The degree of glomerular and peritubular capillary loss in models of progressive renal disease correlates with the severity of glomerulosclerosis and interstitial fibrosis. The aim of our study was to analyze the association of vWF, CD31 and CD34 immunoreactivity with the morphological indices of glomerular sclerosis, interstitial fibrosis, activity and chronicity in GN. A cross-sectional study of 22 patients with GN was conducted. Conventional stains (hematoxylin-eosin, periodic acid Schiff and Trichrome Gömöri stains) and immunohistochemistry (vWF, CD31 and CD34) were employed on kidney biopsies. Activity and chronicity of GN, as well as glomerular segmental sclerosis and interstitial fibrosis, were evaluated according to a scoring system initially used for lupus nephritis and antineutrophil-cytoplasmic-antibody-associated vasculitis. Immunohistochemistry was assessed using a semi-quantitative score. Statistical analysis was performed using EpiInfo 6.04. The mean patient age was 46.68±14.09; 14 patients were male, and eight were female. Performing Spearman`s rank correlation test, no correlation was found between each marker and glomerular segmental sclerosis, interstitial fibrosis, activity and chronicity, which suggests a loss of these markers and microvasculature involvement.

Hematopoietic stem cells have a remarkable plastic capacity, which allows them to differentiate into various cells, such as immune cells, nervous cells, muscle cells, bone and cartilaginous cells. The aim of this study was to show the capacity of stem cells to differentiate into endothelial cells, in culture, after addition of endothelial cells growth suplement (ECGS). We also compared the behavior of these cells with that of endothelial cells obtained from human umbilical vein (HUVEC). CD34+ cells obtained by immunomagnetic separation from human umbilical cord and placental blood were used. After 12‐15 days of culture in a medium containing ECGS, the cells showed morphological changes characteristic to endothelial cells and immunocytochemical analysis revealed the presence of CD31 surface antigen and von Willebrand factor. The flow‐cytometric analysis of endothelial cells adhesion molecules (ECAM) showed that endothelial cells derived from CD34+ cells expressed CD54/ICAM‐1 9.65 ± 0.2% and CD106/VCAM 7.73±0.3%, values similar to those expressed by HUVECs. After TNF incubation, ECAM expression increased only in HUVECs. These data demonstrate that a fraction of circulating CD34+ cells may develop some endothelial cell characteristics when cultured with ECGS, but they are functionally different from HUVECs.

Aristolochic acid (AA) has, in the last decade, become widely promoted as the cause of the Balkan endemic nephropathy and associated renal or urothelial tumours, although without substantial focal evidence of the quantitative dietary exposure via bread in specific households in hyperendemic villages. Occasional ethnobotanical use of Aristolochia clematitis might be a source of AA, and Pliocene lignite contamination of well-water is also a putative health risk factor. The aim of this study was two-fold: to verify if extracts of A. clematitis and Pliocene, or AA by itself, could induce the development of renal or urothelial tumours, and to test the utility of the ribosomal protein p-S6 to identify preneoplastic transformation. Rats were given extracts of A. clematitis in drinking water or AA I, by gavage. After seven months, renal morphology was studied using conventional haematoxylin and eosin and immunohistochemistry for ribosomal p-S6 protein. Plant extracts (cumulative AA approximately 1.8 g/kg b.w.) were tolerated and caused no gross pathology or renal histopathological change, with only faint diffuse p-S6 protein (except in the papilla) as in controls. Cumulative AA I (150 mg/kg b.w. given over 3 days) was also tolerated for seven months by all recipients, without gross pathology or kidney tumours. However, p-S6 protein over-expression was consistent particularly within the renal papilla. In one case given AA I, intense p-S6 protein staining of a proximal tubule fragment crucially matched the pre-neoplastic histology in an adjacent kidney section. We briefly discuss these findings, which compound uncertainty concerning the cause of the renal or upper urinary tract tumours of the Balkan endemic nephropathy.