Biomedical subjects

K W Kim

Publications and source records attributed to K W Kim.

At least 163 records · Page 9Linked to original sources

PubMed1999

Fas is expressed in murine pancreatic islet cells and an insulinoma cell line but does not mediate their apoptosis in vitro.

CD4+ lymphocytes are the most important effector cells in autoimmune diabetes of NOD mice, although some role of CD8+ T cells has been demonstrated. However, it is unknown how CD4+ lymphocytes are able to destroy pancreatic beta-cells that do not express MHC (major histocompatibility complex) class II molecules. Apoptotic cell death mediated by an interaction of Fas with Fas ligand (FasL) could be a mechanism by which MHC class II-negative pancreatic beta-cells are destroyed by CD4+ T lymphocytes. We have examined the expression of Fas molecules in pancreatic islet cells, as well as in a NOD-derived mouse insulinoma cell line (MIN6N8). In addition, the role of Fas-mediated apoptosis in pancreatic islet cell death was explored in vitro. Although Fas expression was not detected by flow cytometric analysis, Fas transcripts were demonstrated in MIN6N8 cells and pancreatic islet cells by the sequencing analysis of the cloned reverse transcription polymerase chain reaction products using Fas-specific primers. IFN (interferon)-gamma, tumor necrosis factor-alpha, interleukin-1 and their combinations failed to enhance Fas expression. Unsorted activated splenocytes from diabetic NOD mice had cytotoxic T lymphocyte activity of a small degree against IFN-gamma-treated MIN6N8 cells with FasL upregulation. However, agonistic anti-Fas antibody with or without cycloheximide did not exert cytotoxicity against MIN6N8 cells or pancreatic islets. FasL transfectant cells also did not kill MIN6N8 cells. Our data indicate that pancreatic beta-islet cells express a small amount of Fas molecules but Fas molecules do not mediate apoptosis of islet cells at least in vitro.

Animals↗

PubMed1999

Obstructive jaundice and acute cholangitis due to papillary stenosis.

Papillary stenosis is characterized by fixed fibrosis leading to structural outflow obstruction and it is usually secondary to inflammation and fibrosis from the chronic passage of gallstones, episodes of acute pancreatitis, chronic pancreatitis, sclerosing cholangitis, peptic ulcer disease, and cholesterolosis. However, obstructive jaundice with or without acute cholangitis which leads the physician to suspect the presence of malignancy as a cause is a rare manifestation of papillary stenosis. We report here a case of papillary stenosis presenting with obstructive jaundice and acute cholangitis. The lesion was so difficult to exclude the presence of malignancy preoperatively and intraoperatively that a pylorus-preserving pancreaticoduodenectomy was performed. Histologic examination of the resected specimen revealed fibrosis, adenomatoid ductal hyperplasia, and mild chronic inflammation of the papilla of Vater and distal common bile duct.

Acute Disease↗

PubMed1999

Early homogeneously enhancing hemangioma versus hepatocellular carcinoma: differentiation using quantitative analysis of multiphasic dynamic magnetic resonance imaging.

The aim of this study was to determine the usefulness of quantitative analysis of multiphasic dynamic contrast-enhanced magnetic resonance (MR) imaging in differentiating early homogeneously enhancing hemangiomas from hepatocellular carcinomas (HCCs). Four-phased dynamic MR imaging at 10 sec (first phase of dynamic contrast-enhanced imaging, P1), 35 sec (second phase, P2), 60 sec (third phase, P3) and 300 sec (delay phase, P4) immediately after intravenous administration of 0.1 mmol/kg Gadolinium-DTPA was obtained with 1.5-T unit with breath-hold multisection FLASH (fast low angle-shot) sequence (TR/TE, 113-130 msec/4.1 msec; flip angle, 80 degrees). Thirty-three HCCs and 18 hemangiomas, homogeneously enhanced on P1, were included in the study. The images were evaluated quantitatively (SNR, signal-to-noise ratio; and CNR, contrast- to- noise ratio of lesions). Quantitatively, mean CNR was higher for hemangiomas than for HCCs on all phases, and the difference in CNRs between hemangioma and HCCs was statistically significant on P3 and P4 (p < 0.0001). When the cutoff for CNR was set at a value of 7.00 on P3 and 1.00 on P4, sensitivity, specificity and accuracy were 94.4%, 93.9%, and 94.1% on P3, and 94.4%, 81.8%, and 86.3% on P4, respectively. There was no statistically significant difference in SNRs between HCC and hemangioma. The differential diagnosis between early, homogeneously enhancing hemangiomas and HCCs was more confidently made with CNRs of lesions on P3 and P4 in dynamic contrast-enhanced MR imaging.

Adult↗

PubMed1999

Drug-resistant pulmonary tuberculosis in a tertiary referral hospital in Korea.

OBJECTIVES: To estimate the resistance rate and to correlate the clinical characteristics of resistant tuberculosis with the patients of pulmonary tuberculosis who were referred to the university hospital. METHODS: We prospectively performed sensitivity tests for all patients who were diagnosed as active tuberculosis by sputum smear or sputum culture from January, 1995 to June, 1996. Patients profile, previous treatment history, patterns of drug resistance, initial chest films and other clinical findings were analysed. RESULTS: Overall, 24(26.0%) of the 92 patients had resistance to at least one drug and 8(8.6%) had resistance to isoniazid(INH) and rifampin(RFP). Among the 66 patients without previous tuberculosis therapy, 11(16.6%) were drug-resistant and 2(3.0%) were multi-drug resistant. Among the 26 patients with previous therapy, 13(50.0%) were drug-resistant and 6(23.0%) were multi-drug resistant. For all 92, resistance to INH was most common (19.5%), followed by RFP (9.7%) and ethambutol (9.7%). Drug resistance was significantly high in previously treated patients and in cavity-positive patients. Treatment failure was also high in previously treated patients with resistant tuberculosis. In patients with primary resistance, treatment failure was not observed. CONCLUSION: Sensitivity tests are strongly recommended in all culture positive patients with previous therapy but, in patients with primary resistance, sensitivity tests are not required. Proper combination chemotherapy should be given under careful surveillance.

Adolescent↗

PubMed1998

Ca2+ influx through carbachol-activated non-selective cation channels in guinea-pig gastric myocytes.

1. Ca2+ microfluorometry (100 microM K5 fura-2) and the voltage-clamp technique were combined to study the effect of carbachol (CCh, 50 microM) in inducing currents (ICCh) through non-selective cation channels (NSCCCh) and increments in global cytosolic Ca2+ concentration (Delta[Ca2+]c). 2. In Na+-containing bath solution, ICCh fell from an initial phasic to a subsequent small (5 %) tonic component; Delta[Ca2+]c fell to zero. Tonic ICCh and [Ca2+]c became prominent after substitution of extracellular 140 mM Na+ by 140 mM Cs+. Tonic ICCh and Delta[Ca2+]c were insensitive to intracellular heparin (3 mg ml-1) and ryanodine (4 microM), i.e. they did not depend on Ca2+ release from sarcoplasmic reticulum (SR). 3. Single channel currents of NSCCCh could be resolved in whole-cell recordings. Substitution of Na+ by Cs+ increased NSCCCh activity by one order of magnitude and slope conductance from 22 to 30 pS. Extracellular quinidine (3 microM) reversibly blocked the NSCCCh activity. 4. Both tonic ICCh and tonic Delta[Ca2+]c (a) followed a similar time course of activation, desensitization and facilitation, (b) were reversibly blocked by 3 microM quinidine, and (c) persisted upon block of SR Ca2+ release. 5. A Ca2+ fractional current of tonic ICCh (fCa) of 0.009 was calculated by comparing the ratio Delta[Ca2+]c (corrected for simultaneous Ca2+ redistribution) over ICCh with depolarization-induced *Delta[Ca2+]c (Delta[Ca2+]c calculated from ICa induced by a 400 ms depolarization from -60 to 0 mV at 2 mM [Ca2+]o, 145 mM [Cs+]o) over ICa. fCa was 0.023 at [Ca2+]o = 4 mM. 6. With 110 mM extracellular CaCl2 and 145 mM intracellular CsCl, ICCh reversed at +19.5 mV suggesting a permeability ratio PCa/PCs of 2.8. 7. We conclude that Ca2+ influx through NSCCCh under physiological [Ca2+]o could induce Delta[Ca2+]c. The fCa was, however, much smaller than the one calculated from the reversal potential.

Algorithms↗

PubMed1998

Activation of c-jun N-terminal kinase/stress-activated protein kinase and the decreased ratio of Bcl-2 to Bax are associated with the auto-oxidized dopamine-induced apoptosis in PC12 cells.

Current concepts of the pathogenesis of Parkinson's disease center on the formation of reactive oxygen species (ROS). Dopamine is one of the major sources of ROS. In this study, the molecular events during the dopamine-induced apoptosis in PC-12 cells were studied using auto-oxidized dopamine. Auto-oxidized-dopamine induced DNA fragmentation and activation of c-jun N-terminal kinase (JNK)/stress-activated protein kinase (SAPK) faster and stronger than dopamine. Furthermore, N-acetylcysteine, an antioxidant, prevented the auto-oxidized dopamine-induced JNK/SAPK activation and DNA fragmentation. Meanwhile, Bcl-2 started to decrease after onset of apoptosis, and Bax was increased up to beginning of apoptosis, and thereafter decreased. Therefore, these results suggested that activation of JNK/SAPK and the decreased ratio of antiapoptotic Bcl-2 to proapoptotic Bax appear to be associated with the dopamine-induced apoptosis.

Acetylcysteine↗

PubMed1998

Activation of NF-kappaB mediates the PMA-induced differentiation of K562 cells.

The role of NF-kappaB during the PMA-induced megakaryocytic differentiation of K562 cells was investigated using K562 cells transfected with each or both subunits of NF-kappaB. The NF-kappaB subunit-transfected cells have shown much higher sensitivity to PMA-induced differentiation than their parental cells. This result was consistent with the findings that PMA-stimulated activities of NF-kappaB were markedly increased in the NF-kappaB subunit-transfected cells in comparison with their parental cells and PMA-induced differentiation was enhanced by pretreatment with IkappaB-alpha antisense oligonucleotide in the NF-kappaB subunit-transfected cells. Meanwhile, there were basically no difference in the basal and PMA-stimulated MAP kinase activities among the parental and NF-kappaB subunit-transfected cells, respectively. However, PMA-induced differentiation was blocked by pretreatment with PD98059, a specific inhibitor of MEK, in both parental and NF-kappaB-transfected cells. Therefore, these results suggest that during the PMA-induced megakaryocytic differentiation of K562 cells, NF-kappaB works downstream of MAP kinase, or that activation of both NF-kappaB and MAP kinase pathways is involved.

Calcium-Calmodulin-Dependent Protein Kinases↗

PubMed1998

Caspase 3 specifically cleaves p21WAF1/CIP1 in the earlier stage of apoptosis in SK-HEP-1 human hepatoma cells.

We report here that p21WAF1/CIP1, an inhibitor of cyclin kinases, underwent proteolytic processing into a smaller fragment, p14, in the early stage of apoptosis in SK-HEP-1 cells. Apoptosis was induced by either staurosporine or ginsenoside Rh2, a ginseng saponin with a dammarane skeleton. Proteolytic processing was the result of caspase-3 activity, which accompanied the early changes in cell morphology and DNA fragmentation. p21WAF1/CIP1 translated in vitro was cleaved into a p14 fragment when incubated with cell extracts obtained from either ginsenoside Rh2-treated or staurosporine-treated cells. Cleavage was equally inhibited in both cases by adding Ac-DEVD-CHO, a specific caspase-3 inhibitor, but not by Ac-YVAD-CHO, a specific caspase-1 inhibitor. Similarly, p21WAF1/CIP1 was efficiently cleaved by recombinant caspase-3, overexpressed in Escherichia coli. Moreover, the endogenous p21WAF1/CIP1 of untreated cell extracts was also cleaved by recombinant caspase 3, as measured by immunoblotting. Mutation analysis allowed identification of two caspase-3 cleavage sites, DHVD112/L and SMTD149/F, which are located within or near the interaction domains for cyclins, Cdks, and proliferating cell nuclear antigen (PCNA). Taken together, these results show that ginsenoside Rh2 and staurosporine increase caspase-3 activity, which in turn directly cleaves p21WAF1/CIP1 during the early stages of apoptosis. We propose that proteolytic cleavage of p21WAF1/CIP1 is a functionally relevant event that allows release of the cyclin/Cdk complex from the p21WAF1/CIP1 inhibitor, resulting in the elevated levels of cyclin/Cdk kinase activity seen in the earlier stage of apoptosis.

Amino Acid Sequence↗

PubMed1998

ATP-sensitive K+ current and its modulation by substance P in gastric myocytes isolated from guinea pig.

To investigate whether ATP-sensitive K+ channels exist in gastric smooth muscle of the guinea pig and whether they are modulated by substance P, we recorded lemakalim-activated K+ currents from freshly isolated cells using the standard whole-cell configuration. With 0.1 mM ATP and 140 mM K+ in the pipette and 90 mM K+ in the bath solution and a holding potential of -80 mV, lemakalim (10 microM) activated a glibenclamide-sensitive inward current with a mean amplitude of -224+/-34 pA. These currents were voltage-independent from -90 to 0 mV and K+-selective. Increasing the intracellular ATP concentrations from 0.1 to 3 mM reduced the lemakalim-activated currents by about five-fold. External barium and cesium inhibited the lemakalim-activated currents in a dose-dependent manner. External tetraethylammonium (10 mM) inhibited the lemakalim-activated currents by 66+/-15%. Bath application of substance P (5 microM) inhibited the lemakalim-activated currents by 53+/-13% and this inhibition was absent when 0.5 mM guanosine 5'-O-(2-thiodiphosphate) (GDPbetaS) was in the pipette. Phorbol 12,13-dibutyrate (PDB) inhibited the lemakalim-activated currents by 71+/-3%. Chelerythrine (1 microM) reduced the substance P-induced inhibition of lemakalim-activated currents by 22.2+/-11.3%. These results suggest the presence of ATP-sensitive K+ channels in gastric smooth muscle and that substance P inhibits ATP-sensitive K+ channels via G-protein through protein kinase C activation.

Action Potentials↗

PubMed1998

Insulin-like growth factor II (IGF-II) secreted from HepG2 human hepatocellular carcinoma cells shows angiogenic activity.

Hepatocellular carcinoma (HCC) is a typical hypervascular tumor. Since insulin-like growth factor II (IGF-II) has been reported to play a significant role in liver regeneration and hepatocarcinogenesis, we initially examined its angiogenic effect on the chorioallantoic membrane (CAM) of 9-day-old chick embryos. We also investigated whether IGF-II secreted from HepG2 human hepatocellular carcinoma cells induces vascularization using the chick embryo CAM. We found that the concentrated conditioned media (CCM) of HepG2 cell culture induced angiogenesis on the CAM. We also identified IGF-II protein in the CCM from HepG2 cells by Western blot analysis. However, CCM from Chang liver cells, which are normal human liver cells and were free of IGF-II, did not induce angiogenesis in the CAM. These results suggest that IGF-II secreted from hepatocellular carcinoma cells may act as an angiogenic factor for the hypervascularization of HCC.

Allantois↗

PubMed1998

Hypoxia-induced apoptosis in human hepatocellular carcinoma cells: a possible involvement of the 6-TG-sensitive protein kinase(s)-dependent signaling pathway.

Apoptosis is a morphologically and biochemically distinct form of cell death which can be triggered by a variety of extracellular agents both during normal developments and in adult pathological states. However, the molecular mechanism of apoptotic cell death due to hypoxia has not been clearly elucidated. In this study, we investigated critical factors involved in hypoxia-induced apoptosis using HepG2, a human hepatocellular carcinoma cell line, as an experimental model. We found that 24 h of exposure of HepG2 cells to hypoxia induced apoptosis, for which de novo protein synthesis was required. Apoptosis was demonstrated by DNA fragmentation and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay. Hypoxia-induced apoptosis was associated with a marked induction of c-jun and c-fos messenger RNAs. Electromobility shift assay showed the increased DNA binding activity of AP-1 during hypoxia, suggesting that AP-1 may be involved in the induction of cell death by acting as a transcriptional regulator. A purine analogue, 6-thioguanine (6-TG), significantly blocked the induction of apoptosis by hypoxia. Moreover, the inductive effect of hypoxia on c-jun expression was also inhibited by 6-TG, whereas the levels of c-fos mRNA and its protein were rather strongly increased. Iodoacetamide (IAA), a non-specific inhibitor of ICE family proteases, also has an inhibitory effect on hypoxia-induced apoptosis. These results suggest that the 6-TG-sensitive protein kinase(s)-dependent signaling pathway may be involved in the apoptotic response of HepG2 cells exposed to hypoxia by increasing the level of c-jun and c-fos and the activity of AP-1 and/or by activating ICE family protease(s).

Apoptosis↗

PubMed1998

Isolation of a novel heat shock protein 70-like gene, pss1+ of Schizosaccharomyces pombe homologous to hsp110/SSE subfamily.

A novel heat shock protein 70 (HSP70) gene, pss1+, of fission yeast, Schizosaccharomyces pombe (S. pombe), has been isolated as a multicopy suppressor of a synthetic lethal mutant of ras1+, which shows severe retardation of growth and aggregation phenotype when the ras1 gene function is absent. The pss1+ gene functionally complements the growth defect of the mutant. Sequence analysis revealed that pss1+ encodes an open reading frame (ORF) of 730amino acids that is homologous to the HSP70 family proteins. The Pss1 has high homology to the Saccharomyces cerevisiae (S. cerevisiae) heat shock protein Sse1p/Msi3p (43% identity) that belongs to the HSP110/SSE subfamily of HSP70. The consensus nucleotide sequence of the heat shock element (HSE) was found in the upstream region of pss1+ gene. The transcript level of pss1+ was moderately abundant during steady-state growth at 25 degrees C and increased a few-fold upon shifting to 42 degrees C. Furthermore, transcription of pss1+ increased in nitrogen-starved conditions. Disruption of the pss1+ gene confers a temperature-sensitive growth phenotype and unexpectedly causes the increase in thermotolerance in S. pombe.

Amino Acid Sequence↗

PubMed1998

Ursolic acid-induced down-regulation of MMP-9 gene is mediated through the nuclear translocation of glucocorticoid receptor in HT1080 human fibrosarcoma cells.

We have previously reported that ursolic acid, a pentacyclic triterpene acid, inhibited the invasion of HT1080 human fibrosarcoma cells by reducing the expression of matrix metalloproteinase-9. Since the chemical structure of ursolic acid is very similar to that of dexamethasone, a synthetic glucocorticoid, we investigated whether ursolic acid acts through the glucocorticoid receptor. The expression of matrix metalloproteinase-9 is thought to be regulated similarly with matrix metalloproteinase-1 and matrix metalloproteinase-3 as containing common 2-O-tetradecanoylphorbol-acetate responsible region, where AP-1 proteins can bind. Dexamethasone has been studied to repress the 2-O-tetradecanoylphorbol-acetate-induced expression of matrix metalloproteinase-1 and matrix metalloproteinase-3 through a glucocorticoid receptor-mediated manner. In Northern blot analysis, we found that ursolic acid reduced the expression of matrix metalloproteinase-1 and matrix metalloproteinase-3 induced by 2-O-tetradecanoylphorbol-acetate. Similarly, ursolic acid down-regulated 2-O-tetradecanoylphorbol-acetate-induction of matrix metalloproteinase-9 gene in the same manner of dexamethasone. RU486, a potent glucocorticoid receptor antagonist, was used for identifying that ursolic acid-induced down-regulation of matrix metalloproteinase-9 expression is mediated by its binding to glucocorticoid receptor. The effect of ursolic acid on the matrix metalloproteinase-9 expression was blocked by RU486, suggesting that ursolic acid acts via a glucocorticoid receptor in the regulation of matrix metalloproteinase-9. Western blot analysis and immunocytochemistry showed that ursolic acid increased glucocorticoid receptor fraction in the nucleus, although it decreased the synthesis of glucocorticoid receptor mRNA. In addition, ursolic acid did not decrease the expression of c-jun and DNA-binding activity of AP-1 to its cognate sequences. Taken together, we suggest that ursolic acid may induce the repression of matrix metalloproteinase-9 by stimulating the nuclear translocation of glucocorticoid receptor, and the translocated glucocorticoid receptor probably down-modulating the trans-activating function of AP-1 to 2-O-tetradecanoylphorbol-acetate responsible element of matrix metalloproteinase-9 promoter region.

Binding Sites↗

PubMed1998

Identification of genes differentially expressed by hypoxia in hepatocellular carcinoma cells.

In order to identify genes differentially expressed under hypoxia (1% O2, 5% CO2, balance N2), we performed mRNA differential display analysis using total RNA extracted from hypoxic and normoxic HepG2, human hepatocellular carcinoma (HCC) cells. Of the differentially expressed genes by hypoxia, some of cDNA fragments were cloned and sequenced. The expression patterns of these clones by hypoxia were confirmed by Northern blot analysis and the quantitative RT-PCR. Down-regulated genes by hypoxia have homology to cDNA sequences encoding cytochrome oxidase subunit II and ADP/ATP translocase, respectively. Up-regulated gene by hypoxia was identified as Homo sapiens oscillin. Moreover, novel genes induced by hypoxia represent partial sequences of cDNAs that have not been reported or functionally identified. Up- or down-regulated expression of these genes in response to hypoxia may contribute to human hepatocarcinogenesis.

Amino Acid Sequence↗

PubMed1998

Insulin-like growth factor II induced by hypoxia may contribute to angiogenesis of human hepatocellular carcinoma.

Insulin-like growth factor II (IGF-II) is highly expressed during hepatocarcinogenesis (P. Schirmacher et al., Cancer Res., 52: 2549-2556, 1992; B. C. Park et al., J. Hepatol., 22: 286-294, 1995). However, the mechanism of its enhanced expression is largely unknown. In this study, we show that IGF-II mRNA levels are increased within six h of exposing human hepatoma cell cultures to hypoxia, suggesting that hypoxia may be a strong stimulus for the induction of IGF-II expression in the process of hepatocarcinogenesis. This finding and the fact that hepatocellular carcinoma (HCC) is a typical hypervascular tumor (M. Mise et al., Hepatology, 23: 455-464, 1996) imply that IGF-II may play an important role in the development of neovascularization of HCC. Here we demonstrate that IGF-II substantially increases vascular endothelial growth factor (VEGF) mRNA and protein levels in a time-dependent manner in human hepatoma cells. The induction of VEGF by IGF-II was additively increased by hypoxia. Moreover, the direct angiogenic activity of IGF-II was observed in the quantitative chick chorioallantoic membrane assay (M. Nguyen et al., Microvasc. Res., 47: 31-40, 1994). These data suggest that IGF-II may be a hypoxia-inducible angiogenic factor in HCC.

Animals↗

PubMed1998

Induction of differentiation of the cultured rat mammary epithelial cells by triterpene acids.

We investigated the effects of triterpene acids (TAs), ursolic acid (UA) and oleanolic acid (OA), on the induction of proliferation and differentiation of normal rat mammary epithelial cells (RMEC) or organoids cultured in Matrigel or primary culture system. To elucidate the effects, we tested their differentiation inducing activities with intercellular communication ability, cell cycle patterns, induction of apoptosis, and morphological differentiation in the three dimensional extracellular culture system. To study the changes of RMEC subpopulation in culture, the cultured cells were isolated, immunostained with peanut lectin (PNA) and anti-Thy-1.1 antibody and then analyzed with flow cytometry. Four different subpopulations, such as PNA and Thy-1.1 negative cells (B-), PNA positive cells (PNA+), Thy-1.1 positive cells (Thy-1.1+), PNA and Thy-1.1 positive cells (B+), were obtained and the size of each subpopulation was changed in culture with time in the presence of TAs. Intercellular communication was observed in culture for 7 days in TAs-treated cells, but not in culture for 4 days with scrape-loading dye transfer technique. G2/M phase cells and the number of apoptotic population were increased in TAs-treated groups in cell cycle analyses. S phase fractions were reduced and the change of G1 phase cells was not observed. The colonies with distinct multicellular structures, such as stellate, ductal, webbed, squamous, lobulo-ductal colonies, were observed in Matrigel culture and the frequencies of each colony were changed in the presence of TAs. These results suggest that UA and OA have differentiation inducing effects on rat mammary epithelial cells in primary or in Matrigel culture.

Animals↗

PubMed1998

In vitro angiogenic activity of Aloe vera gel on calf pulmonary artery endothelial (CPAE) cells.

Angiogenic activity of Aloe vera gel was investigated by in vitro assay. We obtained the most active fraction from dichloromethane extract of Aloe vera gel by partitioning between hexane and 90% aqueous methanol. The most active fraction (F3) increased the proliferation of calf pulmonary artery endothelial (CPAE) cells. In addition, F3 fraction induced CPAE cells to invade type 1 collagen gel and form capillary-like tube through in vitro angiogenesis assay, and increased the invasion of CPAE cells into matrigel through in vitro invasion assay. Furthermore, the effect on the mRNA expression of proteolytic enzymes which are key participants in the regulation of extracellular matrix degradation was investigated by northern blot analysis. F3 fraction enhanced mRNA expression of urokinase-type plasminogen activator (u-PA), matrix metalloproteinase-2 (MMP-2), and membrane-type MMP (MT-MMP) in CPAE cells whereas the expression of plasminogen activator inhibitor-1 (PAI-1) mRNA was not changed.

Aloe↗

PubMed1998

All-trans retinoic acid induced differentiation of rat mammary epithelial cells cultured in serum-free medium.

Retinoids are applied to not only cancer prevention but also cancer chemotherapy by stimulating differentiation of cells. We studied differentiation inducing effect of all-trans retinoic acid (ATRA) by studying proportion of high dense fractions of stem-like cells and the size of S phase fraction in cell cycle. From mammary organoids obtained from 7- to 8-week old F344 female rat mammary gland, we cultured rat mammary epithelial cells (RMEC) and treated physiological doses of 10(-6), 10(-7), and 10(-8) M ATRA from the first day and then cultured for 4, 7, and 14 days. After that, immunostaining was performed using peanut agglutinin (PNA) and anti-Thy-1.1 monoclonal antibody (Thy-1.1) that can be used as markers of differentiation. We separated four different cell subpopulations by flow cytometry: cells negative to both reagents (B-), PNA-positive cells (PNA+), Thy-1.1-positive cells (Thy-1.1+), and cells positive to both reagents (B+). We observed continuous decreases of high dense fractions of stem-like cells (PNA+ subpopulations) for 14 days and as much decreases as high doses of ATRA, which were thought to be proportional to doses of ATRA. We labeled RMEC with bromodeoxyuridine and investigated cell cycle fractions that went through S phase. We observed a tendency of decrease of S phase fraction with time in culture, which is thought to be related to continuous decreases of PNA+ subpopulations and inhibitory role of ATRA on cell cycle. These results suggest that physiological doses of ATRA could stimulate differentiation of RMEC and convert stem-like RMEC to differentiated cells in SFM for a relatively long period of 14 days.

Animals↗