Decreased collagen types I and IV, laminin, CK-19 and α-SMA expression after bone marrow cell transplantation in rats with liver fibrosis.

Bone marrow cells have frequently been tested in animal models of liver fibrosis to assess their role in hepatic regeneration. The mononuclear fraction of bone marrow cells is of particular interest, as many studies show that these cells may be beneficial to treat hepatic fibrosis. In this study, we used the bile duct ligation model to induce hepatic fibrosis in an irreversible manner, and rats were treated with bone marrow mononuclear (BMMN) cells after fibrosis was established. Analysis of collagen types I and IV, laminin and α-SMA showed a decreased expression of these proteins in fibrotic livers after 7 days of BMMN cell injection. Moreover, cytokeratin-19 analysis showed a reduction in bile ducts in the BMMN cell-treated group. These results were accompanied by ameliorated levels of hepatic enzymes GPT (Glutamic-pyruvic transaminase), GOT (glutamic-oxaloacetic transaminase) and alkaline phosphatase (AP). Therefore, we showed that BMMN cells decrease hepatic fibrosis by significantly reducing myofibroblast numbers and through reduction of the collagen and laminin-rich extracellular matrix of fibrotic septa and hepatic sinusoids.

Detection and localization of non-HLA-ABC antigenic sites relevant to kidney rejection on endothelial cells.

Immunological rejection of kidney allografts is usually attributed to presensitization to HLA antigens. However, data on HLA identical transplant rejections indicate that non-HLA antigens may also be involved, and it has been suggested that vascular endothelium represents the main target cell. The purpose of the present study is to describe a method of detecting non-HLA antibodies immunocytochemically. We showed the molecular independence between HLA-ABC molecules identified by the monoclonal antibody w6/32, and antigenic sites identified by a kidney rejection patient serum, previously characterized, on cultured endothelial cells isolated from human umbilical cords by collagenase digestion. Single immunofluorescence staining indicated the molecular independence between these antigenic sites, as the first serum showed a granular pattern, diffused throughout the cytoplasm and the other a reticular pattern restricted to the same cytoplasmic region. This result was confirmed by double labeling. Immunoelectronmicroscopy study also confirmed site independence, showing labeling patterns with different intensities and distinct localizations, using 10- and 20-nm colloidal gold particles to reveal HLA-ABC and non-HLA-ABC determinants, respectively. In conclusion, cultured endothelial cells may be used immunocytochemically to detect non-HLA-ABC determinants of antibody reactivity in renal graft recipients, and the indirect immunofluorescence may be the methodology of choice, since it is easy, reliable and low cost.

Do microtubules around the Toxoplasma gondii-containing parasitophorous vacuole in skeletal muscle cells form a barrier for the phagolysosomal fusion?

The intracellular fate of Toxoplasma gondii was studied in primary cultures of skeletal muscle cells (SMC). The labelling of secondary lysosomes with BSA-Au particles showed no phagolysosomal fusion with the vacuole containing the parasite. After internalization of the parasites, the parasitophorous vacuole became involved by closely apposed endoplasmic reticulum (ER) and mitochondria; within 18 h of interaction, microtubules were visualized in association with the parasitophorous vacuole, suggesting that they could form a barrier for the phagolysosomal fusion.

Ultrastructural localization of anionic sites and lectin-binding sites in sarcoid human alveolar macrophages during interaction with T-lymphocytes.

Sarcoidosis alveolitis is caused by an unknown stimulus activating alveolar macrophages (AM) and T-lymphocytes. During antigen presentation, the complex HLA class II molecule/processed peptide, on the surface of sarcoid AM, induces the T-lymphocyte to proliferate. Altered glycosylation patterns of cell surface glycoproteins such as class II molecules in inflammatory states, may enhance the antigen-presenting capability of AM. In order to know if anionic sites and lectin-binding sites take part in the process of antigen presentation by alveolar macrophages, cells obtained from bronchoalveolar lavage of patients with pulmonary sarcoidosis were incubated with cationized ferritin (CF) and colloidal gold complexed lectins (BSL-I-A4; RCA-I; RCA-II; WGA) for 30 min at 4 degrees C. After incubation, the cells were fixed with 4% paraformaldehyde, 2% glutaraldehyde, postfixed, and Epon embedded. The CF particles were uniformly distributed over the entire cell surface of the lymphocyte, and formed clusters on the surface of the macrophage mainly at the adhesion region between the AM and the lymphocytes. We found enhanced binding of BSL-I-A4 by AM, while WGA and RCA were poorly taken up by these cells. Gold-BSL-I-A4 was distributed randomly on the plasma membrane of the AM, and clustered in the adhesion region with lymphocytes. These results suggest that anionic sites and alpha-D-N-acetyl-galactosamine residues labeled with gold-BSL-I-A4 may be involved in the process of antigen presentation by sarcoid alveolar macrophages.

Progenitor cells and TNF-alpha involvement during morphological changes in pancreatic islets of obese mice.

Overnutrition during pregnancy and lactation lend increasing support to the development of obesity and several chronic diseases in adulthood such as type 2 diabetes mellitus, which leads to beta-cell dysfunction and insulin resistance. In this work, we aimed to study the effects of early life overnutrition on the development of obesity, analyzing the morphological changes, expression of TNF-α, and also the stem cell marker CD133 in the pancreatic islets of young and adult mice. Overnutrition during lactation phase was used as an experimental model to induce obesity. The animals were analyzed at 28 and 150 days of age, when pancreata were collected for histological, ultrastructural and western blotting analysis. The results showed that islet hypertrophy is established in obese groups at day 28 and remained until adulthood. CD133+ cells were observed as small cells within pancreatic islets in both control and obese young mice. However, at day 150, these cells were observed only in the islet peripheries and near ducts of the obese group. Furthermore, TNF-α expression in pancreatic islets was increased in both young and adult obese groups when compared to control groups. This work shows interesting data about CD133 receptor and TNF-α roles in the pancreas during obesity development.