Xenogeneic bone marrow transplantation: II. Porcine-specific growth factors enhance porcine bone marrow engraftment in an in vitro primate microenvironment

Abstract: Establishment of mixed bone marrow chimerism in pig-to-primate transplantation, as a means of inducing specific immune tolerance, will require that both immune and nonimmune barriers be overcome. As a preliminary step in evaluating nonimmune barriers in this system, we have developed an in vitro model of engraftment in which long-term culture of porcine bone marrow-derived hematopoietic cells is supported on preformed primate bone marrow stromal layers. In the absence of cytokine supplementation, primate stromal cells were unable to support long-term porcine hematopoiesis in these cultures. Supplementation with porcine Steel Factor was required for long-term maintenance of hematopoietic progenitor cell content and total hematopoietic activity. Addition of porcine IL-3, in combination with porcine Steel Factor, increased long-term progenitor cell content and hematopoietic activity on primate stroma to levels comparable to that obtained in cultures on porcine stroma. The combination of porcine GM-CSF and Steel Factor increased progenitor cell content and hematopoietic activity early in the cultures, but had little effect in long-term cultures. The Steel Factor and IL-3 combination was species-specific in its action in these cultures, as the corresponding human cytokines were unable to effectively support long-term porcine hematopoiesis. Likewise, the combination of porcine cytokines had only minimal effects on long-term bone marrow culture of primate CD34+ cells I on primate stroma.     

Distal Penile Gangrene in a Patient with Chronic Renal Failure

To date, only 10 cases of distal penile gangrene in patients with chronic renal failure have been reported. This rare condition is believed to result from progressive vascular calcification due to secondary hyperparathyroidism in patients with chronic renal failure. We report an additional case of distal penile gangrene in a 41-year-old man who presented with chronic renal disease and pulmonary tuberculosis. Since some authors have emphasized that aggressive surgical treatment in such cases has a significant mortality rate, we took a more conservative approach to treatment.     

Content of HSP70 in rats with hereditary stress-induced arterial hypertension

Sensitivity of normotensive Wistar rats and NISAG rats (with hereditary arterial hypertension) to heat stress is compared at the organism and cell levels. High temperature sensitivity of NISAG rats correlates with a low content of the main heat shock protein HSP70. This relationship can serve as a biochemical marker of predisposition to arterial hypertension. Translated fromByulleten' Eksperimental'noi Biologii i Meditsiny, Vol. 124, No. 8, pp. 171–173, August, 1997     

Evidence for the presence of a type III secretion system in diffusely adhering Escherichia coli (DAEC).

Diffusely adhering Escherichia coli (E. coli) strains (DAEC) represent a potential cause of diarrhoea in infants, and the detection of type three secretion system (TTSS) genes in DAEC would substantiate their pathogenic nature. In this work, four isolates of DAEC, recovered from stools of diarrhoeic children, were analysed by PCR, in order to detect the presence of TTSS genes. Primers targeted to the escC, escJ, escN and escV, some of the most conserved TTSS genes in enteropathogenic and enterohaemorrhagic Escherichia coli (EPEC and EHEC), were used in order to verify the occurrence of homologous genes in our DAEC isolates. By this approach, we were able to characterise DNA fragments corresponding to putative escJ and escN genes in all DAEC isolates. Furthermore, DNA fragments homologous to the escC and escV genes were also amplified from all isolates. Besides the similarity found among the DAEC esc homologues with EPEC and EHEC esc genes, the nucleotide sequence analysis of the flanking regions of the amplified DNA fragments suggests that the putative DAEC esc genes are organised in the same manner as observed in EPEC and in EHEC strains. The results described here provide strong evidence for the presence of a TTSS in the DAEC strains analysed, implicating a pathogenic nature of these isolates.     

A novel two-step method for the formation of tissue-engineered cartilage by mature bovine chondrocytes: the alginate-recovered-chondrocyte (ARC) method.

Most attempts to tissue-engineer cartilage have involved seeding of cultured cells into a biological or synthetic scaffold. We have developed a novel two-step culture approach that makes possible the in vitro formation of cartilaginous-like tissue by mature adult bovine chondrocytes without the aid of a synthetic matrix. The first step consists of culturing chondrocytes under conditions that maintain their rounded shape and their molecular phenotype as assessed by type II collagen and aggrecan production. This step was accomplished by culturing the isolated chondrocytes in alginate beads until the cells have reestablished a proteoglycan-rich cell-associated matrix (CM). The second step consists of culturing the cells with their CM, after recovery from the beads, on a tissue culture insert with a porous membrane. In this study, young adult bovine articular chondrocytes were cultured in alginate beads in the presence of 10% or 20% fetal bovine serum (FBS). After 7 days of culture, the alginate beads were dissolved by incubating the beads for 20 min in sodium citrate buffer, a calcium chelator. Following a brief centrifugation, the cells with their CM were recovered, resuspended in medium containing 10% or 20% FBS and seeded onto a tissue culture insert. After 1 week of culture on the insert, the individual cells with their CM progressively became incorporated into a mass of cartilaginous tissue. Culture with 20% FBS resulted in the best formation of tissues. These tissues, easily recovered from the insert, were then subjected to biochemical and histological analyses. The biochemical results showed that the chondrocytes remain phenotypically stable in the tissues. The de novo tissue has a relatively high ratio of PG/collagen. Histological examination of the tissue revealed it contained a cartilage-like matrix strongly stained with toluidine blue. This scaffold-free system appears ideal to study, in vitro, the development of transplantable cartilaginous tissue.     

Occupational exposure to hazardous substances and risk of cancer in the area of the mouth cavity, oropharynx, hypopharynx and larynx. A case-control study

A case-control study of squamous cell carcinoma of the upper aerodigestive tract conducted in Heidelberg and Giessen (FRG) provided information on occupational factors in 200 patients and 800 controls (adjusted to sex, age and area of living; 4:1 matched design). The number of subjects exposed to wood dusts, organic chemicals, coal products or to cement was significantly elevated in the tumour group. An increased risk for head and neck cancer was observed after exposition to wood dust (RR = 2,2), organic compounds (RR = 2,4), coal products (RR = 2,7) and especially to cement (RR = 4,4). The cancer risk due to cement exposition showed a positive correlation to the duration of exposition and remained significantly elevated after adjustment for alcohol and tobacco consumption.     

Suppression by Trypanosoma brucei rhodesiense of the capacities of human T lymphocytes to express interleukin-2 receptors and proliferate after mitogenic stimulation.

We studied the suppressive effects induced in phytohemagglutinin (PHA)-stimulated human peripheral blood mononuclear cells (PBMC) by purified blood forms of Trypanosoma brucei rhodesiense. The parasite was found to markedly impair lymphocyte proliferation (measured in terms of [3H]thymidine incorporation). The extent of this effect increased with parasite concentration and was not due to mitogen absorption, depletion of medium nutrients, or PBMC killing by the parasite. Significant reductions in interleukin-2 receptor (IL-2R) expression, determined by flow cytometric analysis, were also observed in PHA-stimulated PBMC cultured in the presence of T. b. rhodesiense as evidenced by marked decreases in the surface density of the receptor. Concomitant decreases in the percentage of IL-2R+ cells were recorded in approximately half of the experiments. A discrete, dimly stained subpopulation of IL-2R+ cells were consistently demonstrable whether or not a reduction in the percentage of IL-2R+ cells occurred. Living, but not glutaraldehyde-fixed, parasites suppressed IL-2R expression. In kinetic studies, a low but reproducible level of suppression of IL-2R was demonstrable as early as 6 h after PHA stimulation; the extent of this effect became considerably more pronounced as additional culture time elapsed. Levels of IL-2 biological activity in cocultures of T. b. rhodesiense with PHA-stimulated PBMC were comparable with or higher than those present in control cultures lacking the parasite. Therefore, insufficient levels of this cytokine would be an unlikely explanation for the noted suppression of IL-2R expression and lymphoproliferation. These effects of T. b. rhodesiense could represent an important component of the mechanism by which immunosuppression develops in African sleeping sickness.