SV40-transformed human fibroblasts exhibit a characteristic chromosomal pattern

A karyotype study of seven SV40-transformed human fibroblast cell lines was performed using R-banding. Although large variations existed from line to line and, to a lesser degree, from cell to cell in a given line, many common features were found. The most characteristic were chromosome imbalances. Some of the chromosomes or chromosome segments present in excess were, in decreasing order of frequency, the early replicating X, 12q, 3q, 12p, 19, 1p, and 6p. Losses of other chromosomes and chromosome segments were also frequent and involved 11p, 2p, 6q, 10p, 18, 4q, 8p, 4p, 10q, and 16. These imbalances seemed to correlate with metabolic characteristics, previously described, such as low activities of catalase (11p), superoxide dismutase 2 (6q), and acid phosphatase (2p, 11p) and a high ratio of lactate dehydrogenase B (LDHB, 12p) activity to LDHA (11p) activity.     

Aluminum ions induce DNA synthesis but not cell proliferation in human fibroblasts in vitro

The effect in vitro of aluminum (Al) ions on DNA synthesis and human dermal fibroblast proliferation using [Al] concentrations from 1.85 to 74 μM and incubation periods of 1, 2, 3, 4, and 5 d was assessed. The lowest concentration of Al that exerted a slight positive, although not significant, effect on DNA synthesis was 1.85 μM, after d 3 or 5 of incubation. The stimulating action of Al was more evident and statistically significant from concentrations of 3.7 μM and 2 d exposure onward. This Al-induced effect on [³H] thymidine incorporation into DNA increased in a time-dependent manner as [Al] in the culture medium rose, provoking increments of up to 322% above the control at [Al] 74 μM and 5 d incubation. In contrast, Al salts moderately increased fibroblast division in a continuous manner only from 7.4 to 74 μM after 3 d of incubation. Although significant overall, the minimal and inconstant mitogenic activity of Al differs greatly from and is not parallel to DNA synthesis, which is not clearly related to exposure times or Al concentrations. Abnormalities in Al-induced cellular metabolic processes described herein and their influence on the cell cycle may constitute a toxicity mechanism for human tissues, leading to disease development. Further studies are required to determine whether these findings can be extrapolated to in vivo situations.     

Proteases stimulate proliferation of human fibroblasts.

Incubation of primary human fibroblasts in serum-free medium with small concentrations of thrombin, trypsin or plasmin resulted in manyfold increase in total DNA synthesis and in the number of 3H-thymidine labelled cells. Rise in the frequency of mitoses indicates that the proteases stimulated also cell division. Because proteases induced only a fraction of cells to proliferate increase in the total number of cells remained moderate. Calf, horse and rabbit serum inhibited the growth stimulating effect of trypsin but chicken, dog and monkey serum were permissive. Specific inhibitors of proteases prevented the stimulation of cell proliferation suggesting strongly that proteases act in virtue of their enzymatic activity.     

Intra-tibial injection of human prostate cancer cell line CWR22 elicits osteoblastic response in immunodeficient rats

We investigated the utility of CWR22 human prostate cancer cells for modeling human metastatic prostate cancer, specifically their ability to induce bone formation following intra-tibial injections in the nude rat. Prostate cancer is unique in regard to its tropism for bone and ability to induce new bone formation. In contrast to humans, other mammalian species rarely develop prostatic cancer spontaneously upon aging and do not have the propensity for bone metastasis that is the hallmark of cancer malignancy in men. We chose human prostate cancer cell line CWR22 based on its properties, which closely resemble all of the features that characterize the early stages of prostatic cancer in human patients including slow growth rate, hormone dependence/independence and secretion of prostate-specific antigen. When CWR22 cells were injected directly into the proximal tibia of immunodeficient male rats, both osteoblastic and osteolytic features became evident after 4 to 6 weeks, with elevated levels of serum prostate-specific antigen. However, osteosclerosis dominates the skeletal response to tumor burden. Radiological and histological evidence revealed osteosclerotic lesions with trabeculae of newly formed bone lined by active osteoblasts and surrounded by tumor cells. Toward the end of the 7-week study, osteolytic bone lesions become more evident on X-rays. Paraffin and immunohistochemical evaluations revealed mature bone matrix resorption as evidenced by the presence of many tartrate resistant acid phosphatase positive multinucleated osteoclasts. We conclude that the CWR22 human prostate cell line used in an intra-tibial nude rat model provides a useful system to study mechanisms involved in osteoblastic and osteolytic bony metastases. This type of in vivo model that closely mimics all major features of metastatic disease in humans may provide a critical tool for drug development efforts focused on developing integrated systemic therapy targeting the tumor in its specific primary or/and metastatic microenvironments. In addition to targeting bone marrow stroma, this strategy will help to overcome classical drug resistance seen at the sites of prostate cancer metastasis to bones.     

Correlation between contractility and proliferation in human fibroblasts

The contractile power of human fibroblasts was checked through their life span in vitro, using a plasma clot retraction test. It was found to decline with a pattern analogous to that of the different phases identifiable by the study of the kinetics of proliferation of these cells. The capacity to retract a plasma clot was higher in cells harvested during active growth than in cells harvested in resting phase. The decreased ability to retract during aging becomes apparent when cells are harvested in resting phase. Decreased retractile activity was also observed in postnatal cells as compared with embryonic cells. The results support a correlation between the initiation of DNA synthesis and the turnover of cytoskeletal elements. The data fit our previous results showing that the early proliferative disturbance during cellular senescence consists of a decline in the probability of initiating the division cycle linked to impaired cell attachment and spreading.     

Immunosuppressive factors from human breast carcinoma cell lines that affect initiation of lymphocyte proliferation

Summary Supernatants from two human breast carcinoma cell lines, 734B and 231, have been shown to inhibit lymphocyte activation by mitogens and antigens. This inhibition appears to be specific for lymphocytes or recently stimulated cells, while having no effect on the growth of established cell lines. Studies of the mechanism of inhibition revealed that the factors inhibit lymphocyte activation and that the factors must be present at the initiation of lymphocyte stimulation for inhibition to occur. The supernatants do not inhibit lymphocyte activation by blocking binding of PHA to lymphocytes. Preliminary purification steps have shown that the inhibitory factors present in the tissue culture supernatants are precipitated at 50% ammonium sulfate saturation and their molecular weights are greater than 100 000. The inhibitory capacity of the 734B supernatants was destroyed by heating at 70° C, while the factors present in the 231 supernatants were only partially destroyed by heating to 90° C. The possible mechanism of action of the inhibitory substances released by tumors and their relevance to tumor growth are important to understanding of immune responses to neoplasia.     

Pericycle cell proliferation and lateral root initiation in Arabidopsis

In contrast with other cells generated by the root apical meristem in Arabidopsis, pericycle cells adjacent to the protoxylem poles of the vascular cylinder continue to cycle without interruption during passage through the elongation and differentiation zones. However, only some of the dividing pericycle cells are committed to the asymmetric, formative divisions that give rise to lateral root primordia (LRPs). This was demonstrated by direct observation and mapping of mitotic figures, cell-length measurements, and the histochemical analysis of a cyclin-GUS fusion protein in pericycle cells. The estimated duration of a pericycle cell cycle in the root apical meristem was similar to the interval between cell displacement from the meristem and the initiation of LRP formation. Developmentally controlled LRP initiation occurs early, 3 to 8 mm from the root tip. Thus the first growth control point in lateral root formation is defined by the initiation of primordia in stochastic patterns by cells passing through the elongation and young differentiation zones, up to where lateral roots begin to emerge from the primary root. Therefore, the first growth control point is not restricted to a narrow developmental window. We propose that late LRP initiation is developmentally unrelated to the root apical meristem and is operated by a second growth control point that can be activated by environmental cues. The observation that pericycle cells divide and lateral root primordia form without intervening mitotic quiescence suggests that lateral organ formation in roots and shoots might not be as fundamentally different as previously thought.     

Changes in cell proliferation,but not in vascularisation are characteristic for human endometrium in different reproductive failures - a pilot study

Background  

Reproductive failure, determined as recurrent spontaneous abortions (RSA) or recurrent implantation failure (RIF) in women is not well understood. Several factors, including embryo quality, and cellular and molecular changes in endometrium may contribute to the insufficient feto-maternal interaction resulting in reproductive failure. Prior clinical studies suggest an inadequate endometrial growth and development of the endometrium, leading to a lesser endometrial thickness.     

Effects of ferrous iron and transferrin on cell proliferation of human diploid fibroblasts in serum-free culture

Summary Iron-free RITC 80-7 defined medium was used to examine effects of ferrous iron and transferrin on cell proliferation of human diploid fibroblasts. Both ferrous iron and holotransferrin stimulated cell proliferation in the medium, but apotransferrin did not. When 5 g/l human serum albumin (HSA) was added to the defined medium, excellent growth was obtained under hypoxic conditions, whereas a reduction of cellular growth during the culture periods was observed under aerobic conditions. When ferrous iron was added to the HSA medium alone, the reduction in growth increased in proportion to the concentrations, whereas the addition of transferrin prevented this reduction in a concentration-dependent manner. This suggests that the ferrous iron concentration in media causes a reduction in growth under aerobic conditions and transferrin prevents this reduction because it decreases the ferrous iron concentration. Further, serum albumin seems to be a source of iron in media.     

Bacillus sphaericus binary toxin elicits host cell autophagy as a response to intoxication

Bacillus sphaericus strains that produce the binary toxin (Bin) are highly toxic to Culex and Anopheles mosquitoes, and have been used since the late 1980s as a biopesticide for the control of these vectors of infectious disease agents. The Bin toxin produced by these strains targets mosquito larval midgut epithelial cells where it binds to Cpm1 (Culex pipiens maltase 1) a digestive enzyme, and causes severe intracellular damage, including a dramatic cytoplasmic vacuolation. The intoxication of mammalian epithelial MDCK cells engineered to express Cpm1 mimics the cytopathologies observed in mosquito enterocytes following Bin ingestion: pore formation and vacuolation. In this study we demonstrate that Bin-induced vacuolisation is a transient phenomenon that affects autolysosomes. In addition, we show that this vacuolisation is associated with induction of autophagy in intoxicated cells. Furthermore, we report that after internalization, Bin reaches the recycling endosomes but is not localized either within the vacuolating autolysosomes or within any other degradative compartment. Our observations reveal that Bin elicits autophagy as the cell's response to intoxication while protecting itself from degradation through trafficking towards the recycling pathways.     

Presence of VIP receptors in a human pancreatic adenocarcinoma cell line. Modulation of the cAMP response during cell proliferation

It is known that the human exocrine pancreas responds to secretin stimulation more than does VIP, a structurally related peptide. We looked for the receptors for those polypeptides in a human pancreatic cancer cell line grown in culture and in nude mice. By analysing the cAMP responses and the 125I-VIP binding we found VIP receptors with a KD of 1.5 10(-9) M. Secretin stimulates the adenylate cyclase through the VIP receptor sites with a KD of 1.7. 10(-6) M. We noted also that during cell proliferation in culture there was about a 5 fold increase of the cAMP response to VIP.     

Increased p21(ras) activity in human fibroblasts transduced with survivin enhances cell proliferation

Survivin is critically involved in mitosis and when overexpressed enhances the activity of the Aurora B kinase, a serine-threonine kinase belonging to the family of oncogenic Aurora/IpI1p-related kinases. Both proteins interact with Ras GTPase-activating protein suggesting an impact on the Ras pathway. This study aimed at defining the role of survivin in proliferation and potential transformation of cells. When survivin was overexpressed in normal human lung fibroblasts, the characteristic track lanes of fibroblasts were disturbed and the rate of cell proliferation was increased. An enhanced level of p21(ras) mRNA and protein expression and concomitant rise in levels of activated p21(ras) were observed. Despite increased proliferation cell survival remained dependent on serum and cells were not able to form colonies in soft agar assays. These data suggest that overexpression of survivin increases cell growth but, despite the increase in active p21(ras), is not sufficient to transform primary cells. Yet, in addition to its anti-apoptotic function it might contribute to the accelerated growth of tumour cells by increasing p21(ras) activity.     

The intracellular form of human MAGP1 elicits a complex and specific transcriptional response

Microfibril-associated glycoprotein-1 (MAGP1) is found associated with microfibrils in the extracellular matrix (ECM). In humans, MAGP1 is expressed as two alternatively spliced isoforms: MAGP1A, the extracellular microfibril-associated form; and MAGP1B, an exclusively intracellular isoform derived from the skipping of exon 3. The biological function of MAGP1B is unknown. We performed gene expression profiling to study the cellular response to MAGP1B using whole-genome genechips. We found that MAGP1B specifically induces the expression of genes linked to cell adhesion, motility, metabolism, gene expression, development and signal transduction. Versican, a gene product involved in the structure and functional regulation of the ECM, showed the highest up-regulation in response to MAGP1B. These studies suggest a dual role for MAGP1, with extracellular MAGP1A involved in ECM function, and intracellular MAGP1B modulating the expression of genes that function in cell adhesion, migration and control of ECM deposition.     

Localization of acetylcholinesterase in normal human fibroblasts and in a human fibrosarcoma cell line

A modified histochemical procedure was used to detect specific acetylcholinesterase localization in cultured human cells. Enzymic activity was found in cytoplasm in the perinuclear zone and in the plasma membrane of 20% to 40% of HT-1080 human fibrosarcoma cells. The presence of pseudocholinesterase was excluded using specific substrates and inhibitors.