[Sorafenib regulates vascular endothelial growth factor by runt-related transcription factor-3 to inhibit angiogenesis in hepatocellular carcinoma].

Objective: To investigate the molecular mechanism of sorafenib against hepatocellular carcinoma. Methods: Sorafenib efficacy was screened and verified by the hepatocellular carcinoma patient-derived tumor xenograft (PDX) model. Veterinary B-mode ultrasonography and in vivo confocal laser scanning microscopy were used to observe PDX angiogenesis. Immunohistochemistry was used to observe the expression of proliferation and angiogenesis-related proteins in PDX tissue. Real-time quantitative PCR technology was used to observe the RUNX3 gene in PDX tissues. SPSS 17.0 statistical software was used for statistical analysis. Results: Four cases of PDX were used to screen the efficacy of sorafenib. PDX1 had a significant response to sorafenib, with an inhibition rate of 68.07%. Compared with the control group, sorafenib had significantly inhibited PDX1 relative tumor volume (5.76±2.14 vs. 11.71±2.87, P<0.05). Cell division index (39.50±7.72 vs. 67.10±9.14, P<0.05) and Ki67 expression (288.6±43.40 vs. 531.70±55.60, P<0.05) were significantly decreased. Veterinary B-mode ultrasonography showed evident blood flow signals in PDX1 tumors. In vivo confocal laser scanning microscopy results showed that sorafenib had significantly reduced the total vessel length (1573.00±236.21 vs. 2675.03±162.00, P<0.05) and area (11 145.33±1931.97 vs. 20 105.37±885.93, P<0.05)) of PDX1 tumors. Immunohistochemical results showed that sorafenib had significantly down-regulated the protein expressions of CD34 (27.55±3.76 vs. 45.47±5.57, P<0.05), VEGF (16.33±2.86 vs. 22.77±3.20, P<0.05) and MVD (38.75±6.01 vs. 55.50±8.61, P<0.05). Real-time PCR results showed that sorafenib had significantly up-regulated RUNX3 gene expression (2.14±0.71 vs. 1.00±0.36, P<0.05). However, there was a negative correlation between the expression of RUNX3 gene and the ratio of VEGF-positive cells in sorafenib group (R2=0.509 7). Conclusion: Sorafenib may inhibit the PDX angiogenesis and the growth of hepatocellular carcinoma by regulating the RUNX3-VEGF pathway.

[Construction of apoptosis-stimulating of p53 protein 2 gene knockout mice and its effect on diethylnitrosamine-induced liver cancer].

Objective: To construct apoptosis-stimulating of p53 protein 2 (ASPP2) gene knockout mice using diethylnitrosamine (DEN)-induced liver cancer model to study the biological functions of ASPP2. Methods: The sgRNA oligonucleotides were constructed, and ASPP2 knockout mice were prepared with the CRISPR/Cas9 system. PCR and sequencing methods were used to identify the genotypes of F0 and F1 generations and their progeny. DEN was used to induce ASPP2+/- mice to establish liver cancer model. Results: PCR and sequencing results showed that ASPP2 gene was successfully knocked out in F0 generation mice. The genotype of F1 generation mice was accorded with ASPP2+/- and had obtained stable heredity. The success rate of DEN-induced liver cancer model (7/8 and 3 / 8) of ASPP2 + /-mice obtained by self-hybridization of F1 generation was significantly higher than that of wild-type mice. Conclusion: ASPP2 knockout mice were successfully constructed based on the CRISPR/Cas9 system. The success rate of DEN-induced liver cancer model of ASPP2 knockout mice was significantly higher than that of the wild-type mice.

Effect of tobacco smoke on hydrogen sulfide-induced rat thoracic aorta relaxation.

Levels of hydrogen sulfide (H2S), a gaseous signaling molecule, are reduced in the serum of individuals who smoke. We hypothesized that tobacco smoke influenced smooth muscle relaxation by decreasing H2S levels and this effect could also influence expression of cystathionine γ-lyase (CSE) and sulfonylurea receptor-2 (SUR-2). The aim of this study was to explore the effect of tobacco smoke on H2S-mediated rat thoracic aorta relaxation and its possible mechanism. Thirty-two Sprague-Dawley rats were divided into four groups: control (C) group, short-term smoker (SS) group, mid-term smoker (MS) group, and long-term smoker (LS) group. H2S concentrations in serum, action of H2S on rat aortic vascular relaxation, and expression of CSE and SUR-2 in thoracic aortic smooth muscle were measured. Although there was no significant difference in H2S between the C and the SS groups, concentration of H2S was significantly reduced in both the LS and MS groups compared to control (P<0.01). Furthermore, H2S was significantly lower in the LS than in the MS group (P<0.05). Rat aortic vascular relaxation was lower in all three treatment groups compared to the control, with the most significant decrease observed in the LS group (P<0.05 compared to the MS group). Expression of CSE and SUR-2 was reduced in the LS and MS groups compared to control (P<0.05), with the lowest levels observed in the LS group (P<0.05). Therefore, tobacco smoke reduced expression of CSE and SUR-2 in rat thoracic aorta, which may inhibit H2S production and vascular dilation.

Effect of tobacco smoke on hydrogen sulfide-induced rat thoracic aorta relaxation

Levels of hydrogen sulfide (H2S), a gaseous signaling molecule, are reduced in the serum of individuals who smoke. We hypothesized that tobacco smoke influenced smooth muscle relaxation by decreasing H2S levels and this effect could also influence expression of cystathionine γ-lyase (CSE) and sulfonylurea receptor-2 (SUR-2). The aim of this study was to explore the effect of tobacco smoke on H2S-mediated rat thoracic aorta relaxation and its possible mechanism. Thirty-two Sprague-Dawley rats were divided into four groups: control (C) group, short-term smoker (SS) group, mid-term smoker (MS) group, and long-term smoker (LS) group. H2S concentrations in serum, action of H2S on rat aortic vascular relaxation, and expression of CSE and SUR-2 in thoracic aortic smooth muscle were measured. Although there was no significant difference in H2S between the C and the SS groups, concentration of H2S was significantly reduced in both the LS and MS groups compared to control (P<0.01). Furthermore, H2S was significantly lower in the LS than in the MS group (P<0.05). Rat aortic vascular relaxation was lower in all three treatment groups compared to the control, with the most significant decrease observed in the LS group (P<0.05 compared to the MS group). Expression of CSE and SUR-2 was reduced in the LS and MS groups compared to control (P<0.05), with the lowest levels observed in the LS group (P<0.05). Therefore, tobacco smoke reduced expression of CSE and SUR-2 in rat thoracic aorta, which may inhibit H2S production and vascular dilation.

AMFR gene silencing inhibits the differentiation of porcine preadipocytes.

Our study clarifies the role of the autocrine motility factor receptor (AMFR) gene in porcine preadipocyte differentiation. AMFR-siRNA was transfected into porcine preadipocytes and the preadipocytes were induced to differentiation. Subsequently, qRT-PCR was conducted to examine changes in mRNA expression of a series of genes in porcine preadipocytes, including AMFR, sterol-regulatory element-binding protein-1a (SREBP1a), SREBP2, insulin-induced gene 1 (Insig1), and Insig2. Expression changes in the mRNA of genes regulating adipocyte differentiation were also analyzed using qRT-PCR, including peroxisome proliferator-activated receptor gamma (PPARγ), CCAAT/enhancer-binding protein alpha (C/EBPα), and Kruppel-like factor 2 (KLF2). Western blot analysis was conducted to examine the changes in AMFR protein expression in porcine preadipocytes. Additionally, morphological changes in differentiated porcine preadipocytes were examined by oil red O staining, and changes in optical density (OD) values were measured using an ultraviolet spectrophotometer. At 24 h after transfection with AMFR-siRNA, AMFR mRNA expression significantly reduced (P < 0.01), and AMFR protein expression markedly decreased (P < 0.05). The mRNA expression of SREBP1a, SREBP2, Insig1, and C/EBPα was significantly reduced (P < 0.01), whereas the expression of KLF2 mRNA was significantly elevated (P < 0.01). After induction of preadipocyte differentiation, the number of lipid droplets decreased in the AMFR-silenced group, and the OD value markedly reduced (P < 0.05). In addition, the expression of C/EBPα mRNA significantly decreased (P < 0.05), whereas the expression of KLF2 mRNA considerably increased (P < 0.05). Taken together, silencing of the AMFR gene inhibits the differentiation of porcine preadipocytes.

Effect of pulsed electromagnetic field therapy on the osteogenic and adipogenic differentiation of bone marrow mesenchymal stem cells.

We investigated the effects of pulsed electromagnetic fields (PEMFs) of 20 Hz/2 mT on the osteogenic and adipogenic differentiation of bone marrow stem cells (BMSCs). Sprague Dawley rat BMSCs were isolated and cultured in vitro. The BMSCs of the third passage were obtained and stimulated by PEMFs of 20 Hz/2 mT. The alkaline phosphatase (ALP) activity was measured according to the ALP assay kit manufacturer instructions, the BMSC osteogenic and adipogenic indicators were detected by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR), and oil red O staining was used to observe the adipose-induced adipogenic differentiation of BMSCs. PEMFs of 20 Hz/2 mT significantly promoted the activity of ALP in the BMSCs (P < 0.01) and mRNA expression of osteogenic proteins (osteocalcin and osteopontin). The PEMFs inhibited the expression of adipogenic transcription factors such as adipokines and adipocyte-binding protein-2, and the adipogenic differentiation of BMSCs. PEMFs of 20 Hz/2 mT can promote osteogenic differentiation and inhibit adipogenic differentiation in BMSCs.

Supramolecular host-guest pseudocomb conjugates composed of multiple star polycations tied tunably with a linear polycation backbone for gene transfection.

A series of novel supramolecular pseudocomb polycations (l-PGEA-Ad/CD-PGEAs) were synthesized by tying multiple low-molecular-weight ß-cyclodextrin (CD)-cored, ethanolamine-functionalized poly(glycidyl methacrylate) (PGEA) star polymers (CD-PGEAs) with an adamantine-modified linear PGEA (l-PGEA-Ad) backbone via the host-guest interaction. The pseudocomb carriers were studied in terms of their DNA binding capabilities, cytotoxicities, and gene transfection efficiencies in the HepG2 and HEK293 cell lines. The pseudocomb l-PGEA-Ad/CD-PGEAs exhibited better plasmid DNA-condensing abilities than their counterparts, CD-PGEA and l-PGEA. Meanwhile, the pseudocomb carriers displayed low cytotoxicity, similar to CD-PGEA and l-PGEA. Moreover, the gene transfection efficiencies of the pseudocomb carriers were much higher than those of CD-PGEA and l-PGEA at various PGEA nitrogen/DNA phosphate molar ratios. Such supramolecular preparation of pseudocomb gene carriers could provide a flexible approach for adjusting the structure and functionality of supramolecular polymers via the proper use of non-covalent interactions.

Computer-aided applications of nanoscale smart materials for biomedical applications.

Nanotechnology has the potential to impact the treatment of many diseases that currently plague society: cancer, AIDS, dementia of various kinds and so on. Nanoscale smart materials, such as carbon nanotubes, C(60), dendrimers and cyclodextrins, hold great promise for use in the development of better diagnostics, drug delivery and the alteration of biological function. Although experimentation is being used to explore the potential offered by these materials, it is by its very nature expensive in terms of time, resources and expertise. Insight with respect to the behavior of these materials in the presence of biological entities can be obtained much more rapidly by molecular dynamics simulation. Furthermore, the results of simulation may be used to guide experimentation so that it is much more productive than it might be in the absence of such information. The interactions of several nanoscale structures with biological macromolecules can already be probed effectively using molecular dynamics simulation. The results obtained should form the basis for significant new developments in the treatment of disease.

Improved lentiviral transduction of human mesenchymal stem cells for therapeutic intervention in pancreatic cancer.

Genetic modification of human bone marrow mesenchymal stem cells (MSC) is highly valuable for their exploitation in basic science and therapeutic applications, for example in cancer. We present here a new, fast and easy-to-use method to enrich a functional population of lentiviral (LV)-transduced MSC expressing enhanced green fluorescent protein (eGFP). We replaced the eGFP gene by a fusion gene of puromycin acetyltransferase and eGFP. Upon LV gene transfer and puromycin selection, we quickly obtained a pure transduced MSC population, in which growth, differentiation capacity and migration preferences were not compromised. Furthermore, we are the first to report the migration velocity of MSC among which 30% were moving and velocity of about 15 mum h(-1) was not altered by LV transduction. Manipulated MSC underwent senescence one passage earlier than non-transduced cells, suggesting the use for therapeutic intervention in early passage numbers. Upon tail vein application in nude mice, the majority of LV-transduced MSC could be detected in human orthotopic pancreatic tumor xenografts and to a minor extent in mouse liver, kidney and lung. Together, LV transduction of genes to MSC followed by puromycin selection is a powerful tool for basic research and improves the therapeutic prospects of MSC as vehicles in gene therapy.

Childhood sexual abuse among outpatients attending adult psychiatric outpatient clinics: a case-control study / Abuso sexual infantil entre los pacientes externos que asisten a las clínicas psiquiátricas ambulatorias para adultos: un estudio de caso-control

Only a few studies have focussed on the importance of routine investigation of childhood sexual abuse in outpatients attending adult psychiatric outpatient clinics. The aim of this study is to explore the association between having a history of childhood sexual abuse (CSA) and attending adult Psychiatric Outpatient Clinics in Trinidad. METHODS: This was a case-control study conducted in twelve psychiatric outpatient clinics located throughout Trinidad A questionnaire covering demographic, social, and sexual abuse components was administered by semi-structured interview to 566 participants, of whom 242 were cases, 239 were controls and 85 had incomplete questionnaires. The cases were 242 patients attending psychiatric outpatient clinics in Trinidad and the controls were 239 non-physician staffmembers at the clinics. Results were analyzed using the Stastistical Package for the Social Sciences (SPSS) version 10. RESULTS: Chi-square analyses revealed several significant differences between the cases and control group. Sixty-three (26%) cases and 29 (12.1%) controls experienced CSA (p < 0.000). Twenty-five (39.7%) of the CSA cases had their experiences between the ages of 4 to 8 years and 13 (44.8%) of the CSA controls had their experiences between the ages of 9 to 12 (p < 0.01). Twenty-six (41.3%) of the cases and 3 (10.3%) of the controls had been abused at least 5 times (p < 0.000). Seventeen (58.6%) abused CSA controls reported having been sexually abused as a child only once. CSA with both force and manipulation was reported by 30 (47.6%) CSA cases while 6 (20.7%) CSA controls experienced CSA with force and manipulation (p < 0. 025). The abused CSA cases reported having a smaller social network of 2 persons compared to the abused CSA controls who had a social network of more than 4 persons (p < 0. 05). Of the 92 abused participants, 73.9% were women, and only 52.2% had told someone about the CSA. For the majority of CSA cases and CSA controls, the abuse involved one abuser. CONCLUSION: A positive correlation was established between earlier onset of CSA, repeated abuse (occurring more than 5 times), a limited social network in patients who had CSA and attending adult psychiatric outpatient clinics. Identifying CSA in psychiatric outpatients may lead to early intervention and aid patient management.

Pocos estudios han centrado su atención en la importancia de la investigación de rutina en relación con el abuso sexual infantil en pacientes externos que asisten a las clínicas psiquiátricas ambulatorias para adultos. El objetivo del presente estudio es explorar la relación que existe entre poseer una historia de abuso sexual infantil (ASI) y asistir a las clínicas psiquiátricas ambulatorias para adultos en Trinidad. MÉTODOS: Se trató de un estudio de caso-control realizado en doce clínicas psiquiátricas para pacientes externos, diseminadas por toda Trinidad. Se administró un cuestionario que abarcaba componentes demográficos, sociales y sexuales, mediante entrevistas semi-estructuradas, a 566 participantes, de los cuales 242 fueron casos, 239 fueron controles, y 85 dejaron sus cuestionarios incompletos. Los casos fueron 242 pacientes que asistían a las clínicas externas de psiquiatría, y los controles fueron 239 miembros no médicos del personal en las clínicas. Los resultados fueron analizados usando la versión 10 del Paquete Estadístico para las Ciencias Sociales (SPSS). RESULTADOS: Los análisis de chi-cuadrado revelaron varias diferencias significativas entre los casos y el grupo de control. Sesenta y tres (26%) casos y 29 (12.1%) controles habían experimentado ASI (p < 0.000). Veinticinco (39.7%) de los casos de ASI tuvieron sus experiencias entre las edades de 4 a 18 años y 13 (44.8%) de los controles de ASI tuvieron sus experiencias entre las edades de 9–12 ( p < 0.01). Veintiséis (41.3%) de los casos y 3 (10.3%) de los controles habían sido víctimas de abuso por lo menos 5 veces (p < 0.000). Diecisiete (58.6%) de los controles víctimas de ASI, reportaron haber sido abusados sexualmente cuando niños, una sola vez. ASI con fuerza y manipulación, fue reportado por 30 (47.6%) casos de ASI mientras que 6 (20.7%) controles de ASi, experimentaron ASI con fuerza y manipulación (p < 0.025). Los casos de abuso de ASI, reportaron haber tenido una red social más pequeña de 2 personas, en comparación con los controles de abuso de ASI, que tenían una red social de más de 4 personas (p < 0.05). De los 92 participantes víctimas de abuso, 73.9% eran mujeres, y sólo 52.2% le habían contado a alguien sobre el ASI. Para la mayor parte de los casos de ASI y controles de ASI, en el abuso hubo un solo abusador implicado. CONCLUSIÓN: Se estableció una correlación positiva entre el comienzo más temprano del ASI, la repetición del abuso (que ocurría más de cinco veces), y una limitada red social en pacientes que fueron víctimas de ASI y asistían a las clínicas psiquiátricas ambulatorias. La identificación de ASI en pacientes psiquiátricos externos, puede conducir a una temprana intervención y tratamiento de ayuda al paciente.

Cycloheximide blocks TGF-beta1-induced apoptosis in murine hepatocytes.

AIM: To study the mechanism of transforming growth factor beta1-induced apoptosis in cultured hepatocytes. METHODS: DNA fragmentation and fluorescent microscopy were used to characterize cell apoptosis. Crystal violet staining was used to assess cell viability. Immunoblotting was used to detect Tak1, p53, and Bax. Dual luciferase assay was used to determine TGF-beta1-induced gene expression. Thin layer chromatography was used to examine ceramide level in AML12 cells. RESULTS: In response to TGF-beta1 treatment, AML12 cells exhibited typical chang es, which was characteristic of apoptosis, such as condensation of chromatin, disintegration of nuclei, and DNA fragmentation. TGF-beta1-induced apoptosis in AML12 cells was completely blocked in the presence of cycloheximide. The inhibitory effect of cycloheximide was accompanied with down-regulation of Tak1 expression and TGF-beta1-induced PAI-1 expression. TGF-beta1 induced p53 expression but not Bax. No increase of ceramide was observed in TGF-beta1-induced apoptosis. CONCLUSION: TGF-beta1-induced apoptosis requires TGF-beta1-induced gene expression.

Propranolol increases phosphatidic acid level via activation of phospholipase D.

AIM: To investigate the propranolol-induced phospholipase D (PLD) activity, its contribution to the increase in the level of phosphatidic acid, and the role of protein kinase C (PKC) in this event. METHODS: A combination of [3H]-myristate labeling, transphosphatidylation reaction, lipid extraction, and thin layer chromatography was used to measure the PLD activity. PKC inhibitors and prolonged phorbol-12-myristate-13-acetate (PMA) treatment were used to study the involvement of PKC in propranolol-induced PLD activation. Immunoblotting was used to detect the intracellular levels of PKC. RESULTS: Treatment of A-549 cells with propranolol in the presence of butanol, resulted in the rapid activation of PLD. Propranolol induced the formation of phosphatidylbutanol (PBut), a unique product of PLD, at the expense of phosphatidic acid (PA) formation. Pretreatment of cells with PKC inhibitors Ro-31-8220, staurosporine, and rottlerin increased the propranolol-induced PLD. Down-regulation of PKC by prolonged treatment of cells with PMA also potentiated the propranolol-induced PLD activity. CONCLUSION: Propranolol induces rapid activation of PLD activity, which results in the increase in intracellular level of PA. The data also indicate that propranolol-induced PLD activity could be negatively regulated by PKC.

The involvement of p38 MAPK in transforming growth factor beta1-induced apoptosis in murine hepatocytes.

We reported in this manuscript that TGF-beta1 induces apoptosis in AML12 murine hepatocytes, which is associated with the activation of p38 MAPK signaling pathway. SB202190, a specific inhibitor of p38 MAPK, strongly inhibited the TGF-beta1-induced apoptosis and PAI-1 promoter activity. Treatment of cells with TGF-beta1 activates p38. Furthermore, over-expression of dominant negative mutant p38 also reduced the TGF-beta1-induced apoptosis. The data indicate that the activation of p38 is involved in TGF-beta1-mediated gene expression and apoptosis.

Boundary detection of retinoblastoma tumors with neural networks.

Consistent and accurate measurement of retinoblastoma tumors is of important clinical value for treatment management. This paper presents an algorithm for the determination of retinoblastoma (RB) tumor to assist in the determination of tumor volume changes throughout treatment periods. The result of the development of a neural network approach for the analysis of three-dimensional ultrasound images shows that it is possible to identify retinoblastoma tumors and accurately determine the front and back boundary of the tumor. The algorithm used was a soft competitive learning network with two inputs. The outputs of the network identify the eye, the tumor, and the back of the eye.

Elimination pathways of [14C]losoxantrone in four cancer patients.

Losoxantrone is an anthrapyrazole derivative in Phase III development in the U.S. for solid tumors, notably breast cancer. To obtain information on the routes of elimination of the drug, a study was conducted in four patients with advanced solid tumors, which involved intravenous administration of 100 microCi of [14C]losoxantrone for a total dose of 50 mg/m(2) during the first course of losoxantrone therapy. Blood, urine, and feces were collected for up to 2 weeks and were analyzed for total radioactivity and parent drug. In addition, feces were profiled for the presence of metabolites. Plasma concentrations of total radioactivity exhibited a temporal pattern similar to the parent drug. Combined recovery of administered total radioactivity from urine and feces was 70% with the majority (87%) of this radioactivity excreted in the feces, presumably via biliary excretion. Feces extracts were profiled for metabolites using a high-performance liquid chromatography method developed to separate synthetic standards of previously identified human urinary metabolites. Only intact losoxantrone was found in the feces. About 9% of the dose was excreted in the urine, primarily during the first 24 h and mostly in the form of parent compound. Collectively, these data indicate that fecal excretion of unmetabolized drug via biliary and/or intestinal excretion is the primary pathway of intravenously administered losoxantrone elimination in cancer patients with refractory solid tumors.

Regulation of phospholipase D activity and ceramide production in daunorubicin-induced apoptosis in A-431 cells.

We demonstrated here that daunorubicin induced apoptosis in A-431 cells, a human epidermoid carcinoma cell line. Treatment of cells with daunorubicin induced chromatin condensation, nuclear fragmentation, internucleosomal DNA degradation, and the proteolytic cleavage of PKC-delta and poly(ADP-ribose) polymerase in A-431 cells. Daunorubicin, as well as sphingomyelinase (SMase) and the exogenous cell-permeable ceramide analogue C(2)-ceramide, inhibited phospholipase D activity stimulated by phorbol 12-myristate 13-acetate or epidermal growth factor (EGF). Like ceramide, daunorubicin also decreased EGF-induced diacylglycerol generation. However, no increase in ceramide level was observed in daunorubicin-induced apoptosis in A-431 cells. Moreover, treatment of A-431 cells with exogenous cell-permeable C(2)-ceramide or SMase did not induce apoptosis. These results indicate that daunorubicin induces apoptosis in A-431 cells via a mechanism that does not involve increased ceramide formation.

Inhibition of actin-myosin subfragment 1 ATPase activity by troponin I and IC: relationship to the thin filament states of muscle.

Troponin I (TnI) is the component of the troponin complex that inhibits actomyosin ATPase activity, and Ca(2+) binding to the troponin C (TnC) component reverses the inhibition. Effects of the binding of TnI and the TnI-TnC (TnIC) complex to actin-tropomyosin (actinTm) on ATPase and on the binding kinetics of myosin subfragment 1 (S1) were studied to clarify the mechanism of the inhibition. TnI and TnIC in the absence of Ca(2+) bind to actinTm and inhibit ATPase to similar levels with a stoichiometry of one TnI or one TnIC per one Tm and seven actin subunits. TnI also binds to actinTmTn in the presence of Ca(2+) with a stoichiometry and inhibition constant similar to those for the binding to actinTm of TnI and Tn in the absence of Ca(2+). Thus, in the presence of Ca(2+), the intrinsic TnI which is released from its binding site on actinTm does not interfere with the binding of an extra molecule of TnI to actinTmTn. The rate of S1 binding to actinTmTnI and to actinTmTnTnI in the presence of Ca(2+) was inhibited to the same extent as upon removal of Ca(2+) from actinTmTn. These studies show that TnI inhibits ATPase by the same mechanism as Tn in the absence of Ca(2+), by shifting the thin filament equilibria from the open state to the closed and blocked states.

Activation of phospholipase D activity in transforming growth factor-beta-induced cell growth inhibition.

Cells regulate phospholipase D (PLD) activity in response to numerous extracellular signals. Here, we investigated the involvement of PLD activity in transforming growth factor-beta (TGF-beta1)-mediated growth inhibition of epithelial cells. TGF-beta1 inhibits the growth of MDCK, Mv1Lu, and A-549 cells. In the presence of 0.4% butanol, TGF-beta1 induces an increase in the formation of phosphatidylbutanol, a unique product catalyzed by PLD. TGF-beta1 also induces an increase in phosphatidic acid (PA) level in A-549 and MDCK cells. TGF-beta1 induces an increase in the levels of DAG labeled with [3H]-myristic acid in A-549 and MDCK cells but not in Mv1Lu cells. No increase of DAG was observed in cells prelabeled with [3H]-arachidonic acid. The data presented suggest that PLD activation is involved in the TGF-beta1-induced cell growth inhibition.

A cytotoxic lobane diterpene from the formosan soft coral Sinularia inelegans.

A new cytotoxic lobane diterpene, ineleganene (1), was isolated from the Formosan soft coral Sinularia inelegans. The structure of compound 1 was determined by 1D and 2D spectral analysis.