Biomedical subjects

A Ohtsuru

Publications and source records attributed to A Ohtsuru.

At least 37 records · Page 2Linked to original sources

PubMed1999

Overexpression of insulin-like growth factor-1 (IGF-I) receptor and the invasiveness of cultured keloid fibroblasts.

Keloid is a dermal fibroproliferative tissue of unknown etiology. Protein tyrosine kinases (PTKs) play an important role in the regulation of cell growth and differentiation. Activation of PTK cascades in keloid fibroblasts is thought to be closely linked to abnormal cell proliferation and migration. We determined the expression profile of PTK genes in normal skin and keloid fibroblasts using the homology cloning method with a degenerated primer. Eight PTK genes were expressed among a total of 46 receptor-type clones. The most abundant type of PTK receptors was the platelet-derived growth factor receptor in both fibroblasts. However, insulin-like growth factor-I receptor (IGF-IR) was overexpressed only in keloid-derived fibroblasts (9 of 24). Immunohistochemical analysis confirmed the high expression of IGF-IR in keloid fibroblasts, but not in normal fibroblasts. To examine the functional properties of the IGF-I/IGF-IR pathway, we investigated cell proliferation and invasion activities of both types of fibroblasts. The mitogenic effect of IGF-I on both fibroblasts was very weak compared with serum stimulation. In contrast, the invasive activity of keloid fibroblasts was markedly increased in the presence of IGF-I, and inhibited by a neutralizing antibody against IGF-IR. Our results indicate the involvement of activated IGF-I/IGF-IR in the pathogenesis of keloid by enhancing the invasive activity of fibroblasts.

Adolescent↗

PubMed1999

Stimulation of in vitro angiogenesis by hydrogen peroxide and the relation with ETS-1 in endothelial cells.

The purpose of this study was to examine the effect of hydrogen peroxide (H2O2) on angiogenesis in cultured endothelial cells. Endothelial cells obtained from bovine thoracic aorta (BAECs) were cultured between two layers of collagen type I to measure the tube formation which is a marker for angiogenesis. Addition of H2O2 (0.1-10 microM) to endothelial cells for various periods increased the rate of tube formation. The maximum stimulation of the tube formation was obtained when cells were exposed to 1 microM H2O2 for 30 min, and the enhancement of tube formation was blocked by catalase (10 U/ml). Both proliferation and migration of BAEC which are known to affect angiogenesis, were also stimulated by the addition of H2O2 (0.1 and 1 microM). Thus relatively low concentrations of H2O2 stimulated angiogenesis, proliferation and migration. Ets-1 is a member of the ets gene family of transcription factors, which binds to the ets binding motif in the cis-acting elements and regulates the expression of certain genes such as proteases including urokinase plasminogen activator (u-PA) and matrix metalloproteinase-1 (MMP-1). Interestingly, H2O2 increased the ets-1 mRNA level in BAECs compared with the basal level. The H2O2-stimulated angiogenesis was completely blocked by an ets-1 antisense oligonucleotide, but not by a mismatched oligonucleotide. These findings indicate that low concentrations of H2O2 stimulate angiogenesis in BAECs, and the stimulation mechanisms may partially involve the enhancement of proliferation and migration. Moreover, the H2O2-induced angiogenesis is likely to be mediated by the transcription factor ets-1.

Animals↗

PubMed1999

Expression of the Axl receptor tyrosine kinase in human thyroid carcinoma.

Protein tyrosine kinases (PTKs) play a crucial role in regulating thyroid cell growth and differentiation. The Axl protein is a member of a new family of receptor tyrosine kinases, of which the ligand, Gas6, a protein S-related molecule, recently was proved to be a mitogenic factor for human thyroid cells. To further investigate the involvement of Axl in human thyroid carcinoma, we examined tissues obtained from 81 patients with thyroid carcinomas, 18 with adenomas, and 13 with adenomatous goiters by immunohistochemistry and in situ hybridization. In addition, among the thyroid carcinomas, we compared Axl expression levels with the grade of differentiation and lymph node metastasis of the carcinoma. Axl was expressed faintly in adenomatous goiter and adenomas, but not in normal thyroid tissues. Among the 81 cases of thyroid carcinoma, 70 (86.4%) showed a positive staining for the Axl protein. Immunopositive (+ +) was detected in papillary carcinomas and anaplastic carcinomas. The level of Axl expression, however, had no correlation with the presence of lymph node metastasis in thyroid carcinomas. In situ hybridization also confirmed the presence of axl mRNA in thyroid carcinoma tissues. These findings suggest that Axl expression may be closely involved in human thyroid tumorigenesis.

Adenoma↗

PubMed1999

Involvement of receptor-type tyrosine kinase gene families in cardiac hypertrophy.

OBJECTIVE: The activation of protein tyrosine kinases (PTKs) has been postulated to be involved in cell differentiation and proliferation. To elucidate the involvement of tyrosine kinase genes in normal and pathological conditions, we analysed the expression patterns of receptor-type PTKs in the normal and hypertensive hypertrophied heart in rats. MATERIALS AND METHODS: Hypertrophied and normal rat hearts were obtained from hypertensive rats; deoxycorticosterone acetate (DOCA)-salt and 2 kidney-1 clip (2K-1C), and their sham-operated rats, respectively. A reverse transcription-polymerase chain reaction (RT-PCR) was performed using degenerated primers which were designed from highly conserved regions in the catalytic domains of receptor-type PTKs. The PCR products were ligated into a sequence vector, and subcloned by transforming bacteria. To compare the expression level of these PTK mRNAs in the normal and hypertrophied heart, we performed semi-competetive RT-PCR and immunohistochemical and Western blot analyses. RESULTS: Nucleotide sequencing of approximately 80 clones of PTKs revealed 10 receptor-type, five nonreceptor-type and two unknown types in the rat heart. Tie-2/Tek, Ryk, insulin-like growth factor-I receptor were abundantly expressed in the rat heart as members of receptor-type PTKs. Immunohistochemistry and RT-PCR demonstrated the presence of platelet-derived growth factor (PDGF)-alpha receptor, PDGF-beta receptor and fibroblast growth factor-3 receptor in both normal and hypertrophied hearts. We also confirmed the presence of Flt-1, KDR/FIk-1, and their ligand vascular endothelial growth factor, c-Met and its ligand hepatocyte growth factor (HGF), and Tie-1, Tie-2/Tek by immunohistochemistry and RT-PCR. The coexpression of cardiac HGF and c-Met in hypertrophied hearts, especially in 2K-1 C rats, was induced more intensively than that in DOCA-salt rats. CONCLUSION: These findings suggest that HGF/c-Met interactions may play an important role in cardiac hypertrophy and remodeling, probably as a result of the activation of the local renin-angiotensin system.

Animals↗

PubMed1999

Expression of insulin-like growth factor I messenger ribonucleic acid in developing osteophytes in murine experimental osteoarthritis and in rats inoculated with growth hormone-secreting tumor.

Osteophytes are one of the characteristic features of osteoarthritis and are often found in acromegalic arthropathy. The aim of this study was to investigate insulin-like growth factor I (IGF-I) involvement in osteophyte formation. One percent collagenase solution was injected into murine knee joints as an osteoarthritis model. In a different animal group, GH-secreting tumor cells were inoculated s.c. to the rat thigh as an acromegaly model. A series of osteophyte formation was examined histologically. IGF-I messenger RNA was detected using the in situ hybridization method. Type I IGF receptors were detected immunohistochemically. In the osteoarthritis model, osteophyte formation appeared as synovial or perichondral cell proliferation adjacent to the articular cartilage on day 5, followed by cartilage formation on day 7 and endochondral ossification on day 14. In the acromegaly model, synovial or perichondral cell proliferation was observed 4 weeks after inoculation, followed by osteophyte formation at 8 weeks. In both models, IGF-I messenger RNA and type I IGF receptor were coexpressed by proliferating synovial or perichondral cells, proliferating chondrocytes, and osteoblasts within the developing osteophytes. These results suggest that IGF-I regulated the initiation and development of osteophyte formation in both models in an autocrine and/or paracrine fashion.

Animals↗

PubMed1999

Prop-1 gene expression in human pituitary tumors.

A novel type of pituitary-specific transcription factor, Prophet of Pit-1 (Prop-1) gene (PROP1), expresses in just early embryonic stage in mouse and closely related as a causative gene in combined pituitary hormone deficiency. We studied PROP1 expression to further clarify its correlation with tumorigenesis and biological behavior in human pituitary adenomas. Eighteen pituitary adenomas and three normal pituitary glands were examined for the expression of PROP1 and POU1F1 by using RT-PCR Pituitary adenomas were diagnosed as non-functioning adenomas (n = 11), prolactinomas (n=5), GH-producing adenoma (n = 1) and ACTH-producing adenoma (n = 1). One of non-functioning adenomas was pituitary carcinoma with cerebellar metastasis and CSF dissemination. Our results demonstrated PROP1 expression in all pituitary tumors examined, in contrast, POU1F1 was detected in 14 of pituitary tumors. PROP1 was also expressed in normal pituitary gland, however, it was not in normal brain tissue, glioblastomas (cell lines and tumor tissues) and meningioma. Our data indicated that PROP1 expression was observed constantly both in the pituitary tumors and normal human adult pituitary tissues, suggesting that PROP1 is an essential transcriptional factor for pituitary specific gene expression in human. Therefore, detection of PROP1 might be a useful indicator for differentiating pituitary adenomas, regardless of their hormonal phenotypes, from other brain tumors.

Adenoma↗

PubMed1999

Effect of 22-oxa-1,25-dihydroxyvitamin D3 on human thyroid cancer cell growth.

To examine whether synthetic vitamin D3 analog, 22-oxa-1,25(OH)2D3 (OCT) has an inhibitory effect on the growth of thyroid carcinoma, we tested the in vitro and in vivo effects of OCT on the growth of a well-differentiated thyroid cancer cell line, NPA. OCT bound to its receptor at the same rate as 1,25(OH)2D3, and inhibited the proliferation of NPA cells in vitro in a dose-dependent manner, similar to that observed with 1,25 (OH)2D3. Northern blot analysis showed that steady-state and fetal bovine serum-stimulated levels of c-myc mRNA were suppressed after 0.5-4 hour treatment with OCT. Transfection studies with the deletion mutants of the 5'-up-stream flanking region of c-myc/chloramphenicol acetyltransferase chimera genes indicated the presence of an OCT responsive element between -410 and -106. Next, we examined OCT effects in implanted NPA tumor cells in nude mice. OCT showed no remarkable hypercalcemic effect compared to 1, 25 (OH2)D3, but OCT and 1, 25 (OH2)D3, had no significant inhibitory effect in vivo after either intra-tumor or intra-peritoneum injection. Our results demonstrate that OCT inhibits the proliferation of well-differentiated thyroid cancer in an in vitro system associated with the suppression of c-myc mRNA, but this inhibitory effect was not reproducible in in vivo model.

Animals↗

PubMed1999

Transforming growth factor-beta stimulates articular chondrocyte cell growth through p44/42 MAP kinase (ERK) activation.

Transforming growth factor-beta1 (TGF-beta1) stimulates articular chondrocyte cell proliferation and extracellular matrix formation. We reported previously that immediate and transient expression of c-fos mRNA through protein kinase C activation is required for the mitogenic effect of TGF-beta1 on cultured rat articular chondrocytes (CRAC). In gel kinase assays using myelin basic protein (MBP) showed that total cell lysates from cells treated with TGF-beta1 caused rapid phosphorylation of MBP, which suggests the involvement of mitogen-activated protein kinase (MAPK) activation. To identify specific MAPK pathways activated by TGF-beta1, we performed in vitro kinase assays using specific substrates. TGF-beta1 induced a rapid activation of extracellular signal regulated kinase (ERK) with a peak at 5 min, which decreased to basal levels within 240 min after TGF-beta1 stimulation. In contrast, the c-jun N-terminal kinase activity increased only about 2.5-fold after 240 min of stimulation and p38 MAPK activity did not change significantly. ERK activation by TGF-beta1 was also confirmed by in vivo phosphorylation assays of Elk1. However, a specific MEK1 inhibitor, PD98059, significantly decreased TGF-beta1 induced Elk1 phosphorylation in a dose-dependent manner. Furthermore, PD98059 reduced the TGF-beta1-induced cell growth by 40%. These results indicate that TGF-beta1 specifically activates MEK1 and subsequent ERK pathways in CRAC, and that the activation of this MAPK pathway plays a role in the mitogenic response to TGF-beta1.

Animals↗

PubMed1999

Involvement of parathyroid hormone-related peptide in cell proliferation activity of human uterine leiomyomas.

Uterine leiomyomas develop from uterine smooth muscle cells, which are known to be regulated by estrogen and other growth factors. The purpose of this study was to investigate the role of expression of parathyroid hormone related-peptide (PTHrP) in the growth of uterine leiomyomas treated or untreated with gonadotropin-releasing hormone agonist (GnRH-a). Thirty-nine leiomyoma tissues were obtained from 36 patients who had been treated with GnRH-a (n=10) or without GnRH-a (n=29). The intensity of PTHrP immunostaining was categorized into three grades; "negative", "weakly positive", and "positive". Leiomyoma cell growth was estimated by the proliferating cell nuclear antigen (PCNA) labeling index (LI) with an image analyser. We also investigated the correlation between PTHrP expression and cell proliferation or histopathological findings. In the GnRH-a-untreated group, LI of the PTHrP "positive" group was significantly higher than that of the PTHrP "negative" group, but the intensity of PTHrP immunostaining did not correlate with LI in the GnRH-a-treated group. PTHrP expression did not correlate with histological findings or clinical parameters (age and phase of menstrual cycle) in either the GnRH-a-treated or the -untreated group. In addition, the expression of mRNA for PTHrP and its receptor was detected in leiomyomas by reverse transcriptase-polymerase chain reaction (RT-PCR). Our results indicate that the expression of PTHrP in leiomyomas correlated positively with cell growth in the GnRH-a-untreated group, suggesting that PTHrP may act as a local cell growth modifier in an autocrine/paracrine fashion on uterine leiomyomas.

Adult↗

PubMed1999

Expression of the ets-1 proto-oncogene in human thyroid tumor.

The proto-oncogene ets-1 is a transcription factor known to control the expression of a number of genes involved in extracellular matrix remodeling. To elucidate the involvement of ets-1 in human thyroid carcinoma, we examined 68 cases of thyroid carcinoma and 38 cases of benign tumors by immunohistochemical means. ets-1 was not expressed in normal thyroid follicular cells. Among the 69 cases of thyroid carcinoma, 61 (89.7%) showed positive staining for the ets-1 protein. According to histologic classification, 47 (97.9%) of 48 papillary carcinomas, 7 (87.5%) of 8 follicular carcinomas, 3 (60.0%) of 5 medullary carcinomas, and 4 (50.0%) of 8 anaplastic carcinomas showed positive staining of ets-1. Among the benign tumors, staining was positive for ets-1 in 8 (40.0%) of 20 adenomas and 3 (16.7%) of adenomatous goiters. In situ hybridization also confirmed the expression of ets-1 mRNA in thyroid carcinoma. Expression of ets-1 mRNA was seen in three kinds of cultured human thyroid carcinoma cell lines and carcinoma tissue obtained at surgery by reverse transcription-polymerase chain reaction. These findings suggest that ets-1 is deeply involved in thyroid carcinogenesis.

Humans↗

PubMed1998

Impaired induction of heat shock protein implicated in decreased thermotolerance in a temperature-sensitive multinucleated cell line.

Cells of a temperature-sensitive mutant line (tsFT101) derived from a mouse mammary carcinoma cell line (FM3A) become multinucleated at a non-permissive temperature of 39 degrees C because of disturbed cytokinesis. To explore how this relates to thermotolerance, we examined the proliferative activity of, and heat shock protein (HSP) expression in, FM3A and tsFT101 cells cultured at 37 degrees C and 39 degrees C after heat shock pretreatment (15 min exposure at 45 degrees C). FM3A cells developed thermotolerance when cultured at both 37 degrees C and 39 degrees C, but whereas tsFT101 cells developed thermotolerance at 37 degrees C, this was markedly reduced at 39 degrees C. Western blot analysis showed similar degrees of expression of constitutive HSP70 (HSP73) in FM3A and tsFT101 cells after heat shock pretreatment at both 37 degrees C and 39 degrees C. However, expression of inducible HSP70 (HSP72) was reduced in tsFT101 cells at 39 degrees C compared to 37 degrees C and to FM3A cells at both 37 degrees C and 39 degrees C. Heat shock pretreatment activated DNA binding of heat shock transcription factor (HSF) in FM3A cells at 37 degrees C and 39 degrees C, but only at 37 degrees C in tsFT101 cells. These results indicate that (1) multinucleation caused by disturbed cytokinesis increases temperature sensitivity, (2) HSP70 is critical for the development of thermotolerance in both FM3A and tsFT101 cells, and (3) decreased expression of inducible HSP70 parallels deficient development of thermotolerance in tsFT101 cells cultured at a non-permissive temperature.

Animals↗

PubMed1998

Distribution of the parathyroid hormone-related peptide and its receptor in the saccus vasculosus and choroid plexus in the red stingray (Dasyatis akajei: Elasmobranch).

1. The exact role of the parathyroid hormone-related peptide (PTHrP) is not fully understood. We used immunohistochemistry to localize the PTHrP and its receptor in the brain of the red stingray, particularly in the saccus vasculosus (SV) and choroid plexus. 2. Immunoreactive PTHrP and its receptor were detected in the epithelial cells of the SV and the choroid plexus. In addition, the neuronal perikarya in the nucleus of the SV located in the hypothalamus is positive for the PTHrP. 3. No PTHrP-containing neurons were detected in the choroid plexus. Extracts of SV and choroid plexus showed positive reactions against the PTHrP and its receptor antibody in Western blot analysis. 4. High levels of immunoreactive PTHrP were detected in the plasma equivalent to those present in human humoral malignant hypercalcemia. In contrast, the immunoreactive PTHrP concentration in the cerebrospinal fluid was below detectable levels. 5. Our results suggest that the regulation of the PTHrP in the SV differs from that in the choroid plexus in the red stingray.

Animals↗

PubMed1998

Increased proliferative activity of osteoblasts in congenital hemifacial hypertrophy.

Congenital hemifacial hypertrophy is expressed in facial asymmetry resulting from unilateral overgrowth. We examined proliferative activity of primary cultured osteoblasts taken from the hemifacial hypertrophic side of an 18-year-old woman compared with the normal side. Both fetal calf serum and basic fibroblast growth factor stimulated osteoblast DNA synthesis, and cultured osteoblast proliferation was markedly increased in the hypertrophic side. Neutralizing antibody against basic fibroblast growth factor was effective to partially block fetal calf serum-induced DNA synthesis. These results suggest that fibroblast growth factor and its receptor signal transduction axis may be selectively involved in affected osteoblasts, leading to progression of congenital hemifacial hypertrophy.

Adolescent↗

PubMed1998

Therapeutic usefulness of wild-type p53 gene introduction in a p53-null anaplastic thyroid carcinoma cell line.

Anaplastic thyroid carcinomas very often harbor the mutations in the tumor suppressor gene p53. We have previously shown that wild-type (wt) p53 gene introduction led to cell growth arrest, but not apoptosis, in p53-null anaplastic thyroid carcinoma cells. The present studies were designed to evaluate other therapeutic effects of wt-p53 gene introduction on p53-null thyroid carcinoma cells, as chemo- and radiosensitization and inhibition of angiogenesis have also been described recently as additional therapeutic advantages of wt-p53 gene introduction in tumor cells with p53 mutations. A p53-null anaplastic thyroid carcinoma cell line, FRO, and a FRO subline stably expressing a temperature-sensitive (ts) mutant of p53 (p53Val138), tsFRO, were used. ts-p53 functions as mutant and wt at nonpermissive (37 C) and permissive (32 C) temperatures, respectively. tsFRO showed a prolonged cell doubling time compared to parental FRO when cultured at 32 C, but the cell growth rate was similar between FRO and tsFRO at 37 C. The cytotoxic and clonogenic assays demonstrated that although the sensitivity to three different anticancer agents (cisplatin, 5-fluorocytosine, and doxorubicin) was unaltered, radiosensitivity was enhanced in tsFRO compared to FRO at 32 C. Unexpectedly, in studies on angiogenesis, expression levels of vascular endothelial growth factor (an angiogenic factor) messenger ribonucleic acid were similar between FRO and tsFRO, and thrombospondin-1 (an antiangiogenic factor) messenger ribonucleic acid and protein levels were about 2.5-fold lower in tsFRO than FRO at 32 C, although any difference could not be detected in their ability to inhibit in vitro angiogenesis with the culture medium conditioned by tsFRO and FRO at 32 C. These results suggest that p53-defective thyroid carcinomas may benefit from the combination of p53 gene therapy and radiotherapy. However, further study will be necessary to clarify the pathological significance of thrombospondin-1 in angiogenesis and thyroid tumor growth.

Antimetabolites, Antineoplastic↗

PubMed1997

Expression of parathyroid hormone-related peptide (PTHrP) in gastric tumours.

Parathyroid hormone-related peptide (PTHrP) is produced by various neoplasms. It has been suggested that it acts as a cytokine for cell proliferation and tumour progression. The purpose of this study was to evaluate PTHrP expression in gastric cancers by immunohistochemistry. PTHrP was expressed in 71 of 92 (77.2 per cent) gastric adenocarcinomas without humoral hypercalcaemia. In contrast, one case (5 per cent) out of 20 adenomas and none of the background non-neoplastic epithelium showed PTHrP immunoreactivity. In carcinomas, PTHrP immunoreactivity was higher in moderately differentiated adenocarcinomas (21/22; 95-5 per cent) and poorly differentiated adenocarcinomas (34/34; 100 per cent) than in well-differentiated adenocarcinomas (10/23; 43 per cent). Furthermore, PTHrP expression was more intense in the deeply invasive portions than in the mucosal carcinomas. High percentages of metastatic tumour cells in regional lymph nodes were immunopositive. PTHrP mRNA expression was confirmed by in situ hybridization in gastric adenocarcinomas. Reverse transcription-polymerase chain reaction (RT-PCR) studies of normal gastric mucosa and four human gastric cancer cell lines detected PTHrP transcription in NUGC-1 (poorly differentiated) and NUGC-3 (poorly differentiated) but not in normal gastric mucosa, MKN-1 (well differentiated), and KATO-III (signet ring cell). These findings suggest that overexpression of PTHrP may be involved in the malignant transformation and progression of gastric carcinomas.

Adenocarcinoma↗

PubMed1997

Expression of parathyroid hormone-related peptide in human osteoarthritis.

To evaluate the involvement of the expression of parathyroid hormone-related peptide gene in human articular cartilage pathology, we performed immunohistochemical staining and in situ hybridization on specimens of femoral head cartilage obtained from 15 patients with osteoarthritis, 11 with rheumatoid arthritis, and 12 control subjects. Parathyroid hormone-related peptide-positive chondrocytes were observed predominantly in degenerated lesions of osteoarthritic tissue and were less evident in rheumatoid arthritic samples, while the normal cartilage expressed little parathyroid hormone-related peptide. In addition, the level of parathyroid hormone-related peptide expression was clearly dependent on the degree of cartilage degeneration; cartilage tissues with moderate degenerative changes contained more positive chondrocytes compared with mildly or severely degenerated cartilage. In situ hybridization confirmed the localization of parathyroid hormone-related peptide protein and demonstrated intense expression of mRNA of the peptide in osteoarthritic samples. This is the first demonstration of parathyroid hormone-related peptide expression in chondrocytes from pathologic articular cartilage of humans. Our results suggest that parathyroid hormone-related peptide may be involved in the pathophysiology of osteoarthritis.

Aged↗

PubMed1997

Gastric cancer associated with overexpression of parathyroid hormone-related peptide (PTHrP) and PTH/PTHrP receptor in relation to tumor progression.

Parathyroid hormone-related peptide (PTHrP) is involved in cell proliferation in both neoplastic and non-neoplastic tissues. We describe an autopsy case of gastric cancer in a patient who showed serum hypercalcemia and overexpression of PTHrP and PTH/PTHrP receptor in the metastatic tumor cells. The primary gastric tumor was poorly differentiated adenocarcinoma, and multiple metastases were present in the bone, multiple visceral organs, peritoneum, and lymph nodes. PTHrP and its mRNA were detected only in the metastatic tumor cells, but not in primary gastric tumor. PTH/PTHrP receptor was also demonstrated immunohistologically in metastatic tumor cells. This case suggests that the expression of PTHrP is related to tumor progression and the poor prognosis in tumors associated with humoral hypercalcemia.

Humans↗

PubMed1997

Profilin gene expression and regulation in a temperature-sensitive breast cancer cell line: tsFT101.

The temperature-sensitive mutant cells (tsFT101) derived from a mouse mammary carcinoma cell line, FM3A, become multinucleated at a non-permissive temperature of 39 degrees C. To further understand the molecular mechanism of such cytokinetic disturbance, we examined the expression of profilin, the main regulator of the transition of globular actin (G-actin) to filamentous actin (F-actin). RT-PCR analysis of mouse profilin cDNA from tsFT101 showed a point mutation (177 A two head right arrow G) which was a wobble mutation causing no change in the encoded amino acid. The expression level of profilin mRNA was, however, diminished in cultured tsFT101 cells under non-permissive temperatures compared with wild-type FM3A cells in association with multinucleation. A stable transfection of profilin cDNA expression vector to tsFT101 cells prevented multinuclear cell formation when cultured at 39 degrees C. In contrast, antisense profilin cDNA expression vector did not alter multinuclear cell formation. The primary cause of the cytokinetic disturbance of tsFT101 cells may be due to the diminished level of profilin gene expression.

Animals↗