PubMed Health⌕ Search

Biomedical subjects

S Murthy

Publications and source records attributed to S Murthy.

At least 37 records · Page 2Linked to original sources
PubMed2001

Asymmetric pigment dispersion in a patient with the unilateral Adie pupil.

PURPOSE: Pigmentary glaucoma is a bilateral disorder. When it occurs asymmetrically or unilaterally, a cause should be sought because it may help us to understand the pathophysiology of this condition better. We describe a patient with unilateral pigmentary glaucoma and the Adie pupil in the same eye and discuss the possible role of the Adie pupil in the development of the pigmentary glaucoma. METHODS: A case report. A 37-year-old woman presented with asymmetric pigment dispersion resulting in pigmentary glaucoma in the right eye and an unilateral Adie pupil in the same eye. RESULTS: We believe that the dilated Adie pupil in the right eye might have resulted in a decreased amount of relative pupillary block. This, then, might have led to an increased contact between the posterior iris and the zonules, resulting in greater pigment dispersion and trabecular obstruction in the right eye. CONCLUSION: It has been proposed that the pigment is liberated from the iris pigment epithelium because of the mechanical rubbing of the posterior iris against the anterior zonular packets. Our case supports this proposed mechanism of pigment release.

Adult↗
PubMed2001

Contralateral pulmonary artery stenosis after left pneumonectomy.

Contralateral pulmonary artery stenosis is a rare complication following pneumonectomy. When extensive intrapericardial dissection is warranted, one must be wary of this potential complication and take measures to avoid it. Postoperatively, a high index of suspicion must be maintained in a patient with a new onset of right-sided heart failure after intrapericardial pneumonectomy. We discuss intraoperative risk factors, postoperative clinical findings, and our strategy for repair.

Arterial Occlusive Diseases↗
PubMed2001

CEP-1347/KT-7515, an inhibitor of SAPK/JNK pathway activation, promotes survival and blocks multiple events associated with Abeta-induced cortical neuron apoptosis.

Although the mechanism of neuronal death in Alzheimer's disease (AD) has yet to be elucidated, a putative role for c-jun in this process has emerged. Thus, it was of interest to delineate signal transduction pathway(s) which regulate the transcriptional activity of c-jun, and relate these to alternate gene inductions and biochemical processes associated with beta-amyloid (Abeta) treatment. In this regard, the survival promoting activity of CEP-1347, an inhibitor of the stress-activated/c-jun N-terminal (SAPK/JNK) kinase pathway, was evaluated against Abeta-induced cortical neuron death in vitro. Moreover, CEP-1347 was used as a pharmacologic probe to associate multiple biochemical events with Abeta-induced activation of the SAPK/JNK pathway. CEP-1347 promoted survival and blocked Abeta-induced activation of JNK kinase (MKK4, also known as MEK-4, JNKK and SEK1) as well as other downstream events associated with JNK pathway activation. CEP-1347 also blocked Abeta-induction of cyclin D1 and DP5 genes and blocked Abeta-induced increases in cytoplasmic cytochrome c, caspase 3-like activity and calpain activation. The critical time window for cell death blockade by CEP-1347 resided within the peak of Abeta-induced MKK4 activation, thus defining this point as the most upstream event correlated to its survival-promoting activity. Together, these data link the SAPK/JNK pathway and multiple biochemical events associated with Abeta-induced neuronal death and further delineate the point of CEP-1347 interception within this signal transduction cascade.

Amyloid beta-Peptides↗
PubMed2001

The role of mutant Apc in the development of dysplasia and cancer in the mouse model of dextran sulfate sodium-induced colitis.

BACKGROUND & AIMS: Differences in genetic background may play a role in the development of ulcerative colitis (UC)-related neoplasia. Loss of heterozygosity (LOH) of APC has been reported in human UC-associated neoplasia. To investigate the role of genetic differences in UC-associated neoplasia, we compared differences in dextran sodium sulfate (DSS) colitis-associated neoplasia between wild-type C57BL/6J mice (WT-DSS) and C57BL/6J mice with a germline mutation in Apc (Min-DSS). METHODS: DSS colitis was induced in female wild-type and Min mice. Age- and sex-matched non-DSS-treated Mins were also studied. Animals were sacrificed after 1 and 2 cycles of DSS. The cecums and large intestines were studied for numbers of dysplasias/cancers. Dysplasias were studied for LOH of Apc. RESULTS: No WT-DSS, 100% of Min-DSS, and 50% of non-DSS-treated Mins had dysplasia. The mean numbers of lesions per mouse were 0 (WT-DSS), 15.6 and 29.3 (1 and 2 cycles Min-DSS, respectively), 1.2 and 1.9 (age-matched control Min, 1 and 2 cycle equivalents, respectively; P < 0.0002, Min-DSS vs. WT-DSS and non-DSS-treated Min; P = 0.03, Min-DSS 2 cycle vs. Min-DSS 1 cycle). Cancers were seen in 0%, 22%, and 40% of non-DSS Min, Min-DSS-1 cycle, and Min-DSS-2 cycle animals, respectively. LOH of Apc was observed in 90.6% of dysplasias and 6% of nondysplastic mucosa. CONCLUSIONS: A germline mutation in Apc contributes significantly to the development of colitis-associated neoplasia. Colitis markedly accelerates the development of dysplasia and cancer in the Min mouse. Dysplasia in Min-DSS occurs through LOH of Apc.

Adenoma↗
PubMed2001

Ipsilateral rotational autokeratoplasty: an alternative to penetrating keratoplasty in nonprogressive central corneal scars.

PURPOSE: To report the outcome of ipsilateral rotational autokeratoplasty performed for nonprogressive central corneal opacities. METHODS: Medical records of 27 patients who underwent ipsilateral rotational autokeratoplasty between June 1995 and December 1998 were retrospectively reviewed. The etiology of corneal opacity, preoperative visual acuity, and size of the trephine used was noted. Any intraoperative and early and late postoperative complications were noted in all patients. The final outcome, visual acuity, and astigmatism at final correction were analyzed. RESULTS: The main etiology of corneal opacity was trauma (44.4%), followed by resolved keratitis (29.6%). The size of the graft ranged from 6.5 to 9 mm. Fifteen patients underwent autokeratoplasty alone; additional procedures were combined in the remaining 12. Wound leak was the most common postoperative complication, which was seen in seven patients. The mean follow-up was 12.11 months (SD = 11.95 months). Five patients (18.5%) were lost to follow-up. Of the remaining 22, 19 (86.36%) had clear grafts and 3 (13.6%) had failed grafts. The mean astigmatism at final correction was 4.25 diopters cylinder (SD = 2.72). Final visual acuity of 20/80 or better was obtained in 13 cases (59.09%). CONCLUSIONS: Contrary to common belief, ipsilateral rotational autokeratoplasty is a safe and effective surgical procedure. It can be considered as an alternative procedure to penetrating keratoplasty in a scenario in which tissue scarcity exists, as well as in cases that have a high risk of developing immunologic allograft rejection. Further prospective studies with preoperative specular microscopy and postoperative corneal topography will provide greater insight into the usefulness of this procedure.

Adolescent↗
PubMed2001

Gene expression of sterol regulatory element-binding proteins in hamster small intestine.

Gene expression of sterol regulatory element-binding proteins 1a, 1c, and 2 (SREBP-1a, -1c, and -2) and of 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) synthase, HMG-CoA reductase, and the low density lipoprotein (LDL) receptor was examined in hamster small intestine. SREBP-1c transcript predominated over SREBP-1a. mRNA levels for SREBP-1a, -1c, and -2, LDL receptor, and HMG-CoA synthase were highest in jejunum and ileum. Expression of SREBP-1a and SREBP-1c was highest in cells of the upper villus and decreased in cells of the lower villus. Gene expression of SREBP-2 was lowest in cells of the upper villus and increased in cells of the lower villus. Ileal SREBP-2 gene expression was highest in cells of the midvillus. mRNA levels for HMG-CoA synthase and the LDL receptor followed a pattern similar to that of SREBP-2. A positive correlation existed between SREBP-2 gene expression and rates of cholesterol synthesis. Fatty acid synthesis was highest in jejunum and ileum, correlating positively with the expression of SREBP-1c. Sterol influx into intestinal cells decreased mRNA levels of SREBP-2, HMG-CoA reductase, HMG-CoA synthase, and LDL receptor. In ileum, sterol influx decreased gene expression of SREBP-1a and increased expression of SREBP-1c. The results suggest that SREBP-2 regulates cholesterol synthesis in the small intestine. SREBP-1a is a minor transcript and its expression does not correlate with cholesterol-synthesizing activity. SREBP-1c is a major transcript in small intestine and its expression along the length of the gut correlates with fatty acid synthesis. Sterols regulate gene expression of sterol-responsive genes, including SREBP-2, in small intestine. - Field, F. J., E. Born, S. Murthy, and S. N. Mathur. Gene expression of sterol regulatory element-binding proteins in hamster small intestine. J. Lipid Res. 2001. 42: 1;-8.

Animals↗
PubMed2001

Regulation of sterol regulatory element-binding proteins by cholesterol flux in CaCo-2 cells.

The regulation of sterol regulatory element-binding proteins (SREBP) by cholesterol flux was studied in the intestinal cell line CaCo-2. CaCo-2 cells were incubated for 18 h with micelles containing 5 mM taurocholate and 500 microM oleic acid or micelles containing either 200 microM cholesterol or 150 microM lysophosphatidylcholine. In some incubations, an ACAT inhibitor was added or 25-hydroxycholesterol was substituted for cholesterol. The SREBP-1a transcript was 2-fold more abundant than the SREBP-1c transcript. In cells incubated with micelles containing cholesterol, rates of cholesterol synthesis were decreased and rates of esterification were increased. Cholesterol synthesis was decreased further by ACAT inhibition. Cholesterol influx decreased mRNA levels of SREBP-2, HMG-CoA synthase, HMG-CoA reductase, and fatty acid synthase. ACAT inhibition modestly suppressed gene expression further. Neither SREBP-1a nor SREBP-1c mRNA levels were altered by cholesterol. Despite decreases in gene expression of the sterol-responsive genes by cholesterol, the amounts of precursor and mature forms of SREBP-1 and SREBP-2 were not altered. In contrast, if 25-hydroxycholesterol was substituted for cholesterol, both the precursor and mature forms of SREBP-2 were decreased. The polar sterol decreased the mature form of SREBP-1 but the amount of the precursor form was unchanged. In cells incubated with micelles containing lysophosphatidylcholine, which causes cholesterol to efflux from cells, sterol-responsive gene expression was increased. The amounts of precursor and mature forms of SREBP-1 and SREBP-2, however, were not altered. In contrast, if the cells were depleted of cholesterol by incubating them with lovastatin and cyclodextrin, the mature forms of SREBP-1 and SREBP-2 were increased, as were mRNA levels for the sterol-responsive genes. The data would suggest that cholesterol influx/efflux regulates mRNA levels of sterol-responsive genes independently of changes in the amount of mature SREBP. In contrast, 25-hydroxycholesterol influx or cholesterol depletion alters the amount of mature SREBP, leading to the regulation of sterol-responsive gene expression.

CCAAT-Enhancer-Binding Proteins↗
PubMed2000

Tumor necrosis factor-alpha and interleukin-1beta inhibit apolipoprotein B secretion in CaCo-2 cells via the epidermal growth factor receptor signaling pathway.

In inflammatory conditions of the gut, cytokines are released into the mucosa and submucosa propagating and sustaining the inflammatory response. In CaCo-2 cells, we have shown that various inflammatory cytokines interfere with the secretion of lipids, an effect that is likely caused by the release of a ligand to the epidermal growth factor (EGF) receptor. In the present study, the role of the EGF receptor signaling pathway and the effects of the cytokines tumor necrosis factor-alpha (TNF-alpha) and and interleukin 1beta (IL-1beta) on triacylglycerol-rich lipoprotein secretion were investigated. CaCo-2 cells were incubated with oleic acid to enhance triacylglycerol-rich lipoprotein secretion. TNF-alpha and IL-1beta significantly decreased the basolateral secretion of apolipoprotein B (apoB) mass, with IL-1beta being more potent. Tyrphostin, an inhibitor of the EGF receptor intrinsic tryosine kinase, prevented or markedly attenuated the decrease in apoB secretion by TNF-alpha or IL-1beta. Both cytokines increased the phosphorylation of the EGF receptor by 30 min. Moreover, phosphotyrosine immunoblots of the EGF receptor demonstrated an increase in tyrosine residues phosphorylated by 0.5 and 6.5 h. At both these time points, TNF-alpha and IL-1beta also decreased the binding of EGF to its cell surface receptor. At 6.5 h, activation of the EGF receptor was sustained. In contrast, the early activation of the receptor was only transient as receptor phosphorylation and binding of EGF to its receptor returned to basal levels by 2 h. Preventing ligand binding to the EGF receptor by a receptor-blocking antibody attenuated receptor activation observed after 6.5 h. This did not occur at 0.5 h, suggesting that early activation of the EGF receptor was non-ligand-mediated. Similarly, apoB secretion was inhibited by an early non-ligand-mediated process; whereas at the later time, inhibition of apoB secretion was ligand-mediated. Thus, the inflammatory cytokines TNF-alpha and IL-1beta interfere with the secretion of triacylglycerol-rich lipoproteins by both early and delayed signaling events mediated by the EGF receptor signaling pathway.

Apolipoproteins B↗
PubMed2000

Increased permeability in dextran sulphate colitis in rats: time course of development and effect of butyrate.

BACKGROUND: Increased mucosal permeability is an important factor in the genesis of mucosal inflammation in inflammatory bowel disease. This study examined the time course of increased permeability and the effect of butyrate on permeability in experimental colitis in rats. METHODS: Colitis was induced in albino rats by administration of 4% dextran sulphate sodium (DSS) orally for up to 7 days. Rats were killed sequentially after 1-7 days of DSS feeding and compared with control animals. Distal colon sheets, from normal and DSS rats, were mounted in Ussing chambers. Electric resistance and passive permeation of 14C-mannitol were measured over 90 min. In control and 5-day DSS rats additional permeability measurements were made in the presence of butyrate (25 mmol/l) in the bathing solutions. The permeability of the normal distal colon was measured after addition of DSS in vitro. Sections of colon were examined by light microscopy. The viability of colonocytes, from normal and DSS rat colon, was measured by release of lactate dehydrogenase immediately and during a 60-min incubation after isolation. RESULTS: Focal mild inflammation and shedding of epithelium were noted after 2 days of DSS administration; crypt loss with flattened epithelium in adjacent areas after 5 days; and fibrosis after 7 days. Decreased epithelial cell survival after 60 min of incubation was noted after 1 day of DSS administration, whereas decreased viability at the time of isolation was noted after 2 days of DSS administration (viability, 72.7% +/- 1.4%; mean +/- standard error) compared with control (89.3% +/- 0.8%) (P < 0.01). Increased permeability was noted after 1 day of DSS administration. Electric resistance (mu omega/cm2/h) was significantly reduced after 1 day of DSS administration to 85.9 +/- 4.6 (mean +/- standard error) compared with control animals (117.2 +/- 2.2; P < 0.001). Serosa-mucosa flux of mannitol (micromol/cm2/h) was also significantly increased after 1 day of DSS feeding (0.169 +/- 0.01) compared with control (0.061 +/- 0.08) (P < 0.01). Electric resistance and mannitol permeability were significantly returned towards normal by the presence of butyrate. DSS added directly to the bathing solution did not significantly alter the colon permeability in vitro. CONCLUSIONS: Increased mucosal permeability is a very early change in colitis induced by DSS, is accompanied by decreased cell survival, and precedes detectable changes in histology. Reversal of increased mucosal permeability by butyrate may explain its utility in the therapy of inflammatory disease of the colon.

Animals↗
PubMed2000

Dysplasia and cancer in the dextran sulfate sodium mouse colitis model. Relevance to colitis-associated neoplasia in the human: a study of histopathology, B-catenin and p53 expression and the role of inflammation.

Animal models of colitis, which develop dysplasia and cancer similar to human ulcerative colitis are needed to further investigate the dysplasia cancer sequence. This study describes the expression of B-catenin and p53 along with the histopathology and inflammation scores as they relate to dysplasia and cancer in the dextran sulfate sodium (DSS) colitis model. Swiss Webster mice were fed with 5% DSS as follows: group A, four cycles of DSS, 84 days total (1 cycle = 7 days DSS + 14 days H(2)O); group B, four cycles DSS followed by 120 days H(2)O, 204 days total; group C, 7 days DSS followed by 180 days H(2)O, 187 days total; group D, 7 days DSS followed by 90 days H(2)O, 97 days total. The incidences of dysplasia and/or cancer were 15.8, 37.5, 18.1 and 0% in groups A-D, respectively. Dysplasia and/or cancer occurred as flat lesions or as dysplasia-associated lesion or mass (DALM) as observed in the human. Thirty-three percent of cancers had associated dysplasia. Within group A, inflammation scores were significantly higher in animals with dysplasia and/or cancer compared with those without dysplasia and/or cancer (P < 0. 05-P < 0.0001). Inflammation scores were significantly higher in animals with cancers versus those with dysplasia (P < 0.015) and in flat dysplasia and/or cancer versus DALM (P < 0.0042). B-catenin showed translocation from the cell membrane to the cytoplasm and/or nucleus in 100% of DALM and 5.8% of flat dysplasia and/or cancer. A total of 94.2% of flat dysplasia and/or cancer had exclusive cell membrane expression compared with 0% DALM (P < 0.0001). Only 7.4% of dysplasia and/or cancer showed nuclear expression of p53. In colitis-associated dysplasia and/or cancer in the DSS model: (i) histology resembles that in the human; (ii) inflammation plays a significant role in the dysplasia cancer sequence and whether dysplasia and/or cancer grows as a flat lesion or a DALM; (iii) the early molecular pathways are different for flat dysplasia and/or cancer versus DALM, with nuclear/cytoplasmic translocation of B-catenin as an early event in DALM but not flat dysplasia and/or cancer; and (iv) p53 has little or no role in dysplasia and/or cancer. This well characterized model provides an excellent vehicle for studying the roles of inflammation, the molecular events and the role of chemopreventive agents in colitis-associated neoplasia.

Animals↗
PubMed2000

Corneal triple procedure: indications, complications, and outcomes: a developing country scenario.

PURPOSE: We report the indications, complications, and outcomes of 104 corneal triple procedures in our institute. METHODS: Patient records of 104 consecutive cases of corneal triple procedure (penetrating keratoplasty with extracapsular cataract extraction and intraocular lens implantation) performed by experienced corneal surgeons between January 1992 and December 1997 were retrospectively reviewed. Relevant preoperative, operative, and postoperative data were collected. The outcome was assessed by the graft clarity and visual acuity at the last visit. Survival analysis of these grafts was determined by Kaplan-Meier method. RESULTS: Of 104 patients, 70 were men and 34 were women. Mean age of these patients was 48.5+/-17.1 years (range, 1-75 years). Corneal scarring with cataract was the reason for surgery in 69 (66.4%) cases. The intraoperative complications included vitreous upthrust in seven (6.9%) cases and posterior capsular dehiscence in three (2.9%) cases. The most common early postoperative complications were increased intraocular pressure in 19 (18.3%) and increased anterior chamber reaction in 25 (24%) cases. Posterior capsular opacification was seen in 26 (25%), graft reaction in 15 (14.4%), and secondary glaucoma in 15 (14.4%) cases. These formed the important late postoperative complications. The average postoperative follow-up was 23.7+/-17.6 months (range, 1.6-79.4 months); at which time 72% of the grafts remained clear. At last follow-up, 40% of patients had a visual acuity of > or = 20/40. CONCLUSIONS: Corneal scarring with cataract is the most common reason for triple procedure in this part of the world. This is a safe surgical procedure with good graft clarity and reasonable visual recovery.

Adolescent↗
PubMed2000

Effect of CIS on activity in trans of the replication initiator protein of an IncB plasmid.

RepA, the replication initiator protein of the IncB plasmid pMU720, acts preferentially in cis. The cis activity of RepA is thought to be mediated by CIS, a 166-bp region of DNA separating the coding region of repA from the origin of replication (ori) of pMU720. To investigate the trans activity of RepA, the repA gene, without its cognate ori, was cloned on a multicopy plasmid, pSU39. The ori on which RepA acts was cloned on pAM34, a plasmid whose replicon is inactive without induction by isopropyl-beta-D-thiogalactopyranoside (IPTG). Thus, in the absence of IPTG, replication of the pAM34 derivatives was dependent on activation of the cloned ori by RepA produced in trans from the pSU39 derivatives. The effect of CIS, when present either on the RepA-producing or the ori plasmid or both, on the efficiency of replication of the ori plasmid in vivo, was determined. The presence of CIS, in its native position and orientation, on the RepA-producing plasmid reduced the efficiency of replication of the ori plasmid. This inhibitory activity of CIS was sequence specific and involved interaction with the C-terminal 20 to 37 amino acids of RepA. By contrast, CIS had no effect when present on the ori plasmid. Initiation of replication from the ori in trans was independent of transcription into CIS.

DNA Helicases↗
PubMed2000

Five human genes encoding F-box proteins: chromosome mapping and analysis in human tumors.

Members of the F-box protein (Fbp) family are characterized by an approximately 40 amino acid F-box motif. SCF complexes (formed by Skp1, cullin, and one of many Fbps) act as protein-ubiquitin ligases that control the G(1)/S transition of the eukaryotic cell cycle. The substrate specificity of SCF complexes is determined by the presence of different Fbp subunits that recruit specific substrates for ubiquitination. Unchecked degradation of cellular regulatory proteins has been observed in certain tumors and it is possible that deregulated ubiquitin ligases play a role in the altered degradation of cell cycle regulators. We have recently identified a family of human Fbps. As a first step aimed at determining if FBP genes could be involved in human neoplasia, we have mapped the chromosome positions of 5 FBP genes by fluorescence in situ hybridization (FISH) to 10q24 (BTRC alias beta-TRCP/FBW1a), 9q34 (FBXW2 alias FBW2), 13q22 (FBXL3A alias FBL3a), 5p12 (FBXO4 alias FBX4) and 6q25-->q26 (FBXO5 alias FBX5). Since most of these are chromosomal loci frequently altered in tumors, we have screened 42 human tumor cell lines and 48 human tumor samples by Southern hybridization and FISH. While no gross alterations of the genes encoding beta-Trcp/Fbw1a, Fbw2, Fbx4 and Fbx5 were found, heterozygous deletion of the FBXL3A gene was found in four of 13 small cell carcinoma cell lines. This is the first evaluation of genes encoding Fbps in human tumors.

Amino Acid Motifs↗
PubMed2000

Transient effects of long-term leptin supplementation in the prevention of diet-induced obesity in mice.

Low plasma leptin levels have been shown to be associated with the development of obesity in mice as well as in humans. The present study was undertaken to determine if raising plasma leptin levels of obesity-prone C57BL/6J (B6) mice to those seen in obesity-resistant A/J mice would prevent the development of diet-induced obesity. Four-week-old B6 (n = 40) and A/J (n = 10) male mice were weaned onto a low-fat (11% kcal) diet. When the animals weighed 20 g, their diets were changed to a high-fat (HF) diet (58% kcal), and a continuous infusion of leptin (0.4 mg x kg(-1) x day(-1)) or phosphate-buffered saline (control) was started using Alzet minipumps. The A/J mice were not treated but were included to monitor the efficacy of the minipumps in raising plasma leptin in B6 mice. The mice were followed for 12 weeks. Chronic treatment with leptin for 4 weeks raised plasma levels in B6 mice to that of A/J mice. Plasma leptin in B6 control mice remained significantly lower than A/J mice through week 4. By week 8, leptin levels in the B6 control group had risen and were similar to A/J mice. Although there were significant weight differences between B6 treated and B6 control groups for 2-3 weeks after pump implantation, these differences were transient. Ultimately, there were no weight differences between the B6 treated and B6 control groups. There were no differences in plasma glucose between B6 treated and control groups. Plasma insulin values were also not different between the 2 groups. There was no effect of leptin supplementation on locomotor activity or food intake in B6 mice. In summary, this study demonstrates that leptin supplementation in animals that show low plasma leptin levels in response to fat feeding may slow but does not prevent the subsequent development of diet-induced obesity.

Adipose Tissue↗
PubMed1999

Rank-order of potencies for inhibition of the secretion of abeta40 and abeta42 suggests that both are generated by a single gamma-secretase.

The Alzheimer's disease amyloid peptide Abeta has a heterogeneous COOH terminus, as variants 40 and 42 residues long are found in neuritic plaques and are secreted constitutively by cultured cells. The proteolytic activity that liberates the Abeta COOH terminus from the beta-amyloid precursor protein is called gamma-secretase. It could be one protease with dual specificity or two distinct enzymes. By using enzyme-linked immunosorbent assays selective for Abeta40 or Abeta42, we have measured Abeta secretion by a HeLa cell line, and we have examined the dose responses for a panel of five structurally diverse gamma-secretase inhibitors. The inhibitors lowered Abeta and p3 secretion and increased levels of the COOH-terminal 99-residue beta-amyloid precursor protein derivative that is the precursor for Abeta but did not alter secretion of beta-amyloid precursor protein derivatives generated by other secretases, indicating that the inhibitors blocked the gamma-secretase processing step. The dose-dependent inhibition of Abeta42 was unusual, as the compounds elevated Abeta42 secretion at sub-inhibitory doses and then inhibited secretion at higher doses. A compound was identified that elevated Abeta42 secretion at a low concentration without inhibiting Abeta42 or Abeta40 at high concentrations, demonstrating that these phenomena are separable pharmacologically. Using either of two methods, IC50 values for inhibition of Abeta42 and Abeta40 were found to have the same rank-order and fall on a trend line with near-unit slope. These results favor the hypothesis that Abeta variants ending at residue 40 or 42 are generated by a single gamma-secretase.

Amyloid Precursor Protein Secretases↗
PubMed1999

Tyrosine kinase inhibitors as antiproliferative agents against an estrogen-dependent breast cancer cell line in vitro.

BACKGROUND AND OBJECTIVES: Receptor tyrosine kinase (RTK) activation is critical for growth factor-mediated cell proliferation. Blockade of RTK activation inhibits growth factor-induced cell proliferation. A panel of RTK inhibitors (tyrphostins) have been tested and compared for their antiproliferative effects on the hormone-dependent human breast cancer cell line, MCF-7, in vitro. METHODS: MCF-7 cells (10(4)/well) were seeded into 96 well plates and maintained in DMEM with 1% bovine serum albumin (BSA), 200-pg/mL estrogen, or 10% fetal bovine serum. After a defined time interval, the cells were exposed to RTK inhibitors and a non-RTK-inhibitory analog of tyrphostins (0 to 400 microM). After 3 days, the number of viable cells in each well was estimated by an MTT assay and the results expressed as percent of controls. Using a representative tyrphostin, A47, the validity of MTT assay as a measure of cell proliferation was tested by a colony formation assay and by immunostaining with Ki-67 antibodies. RESULTS: MCF-7 cells maintained in DMEM containing 1% BSA without E2 or serum showed a minimal increase in cell number. Supplementation with E2 stimulated cell proliferation in a dose-dependent manner. This E2-mediated growth stimulation was completely inhibited (cytostatic effects) by the epidermal growth factor receptor (EGFR)-selective tyrphostins A47, B48, RG13022, and B50. These same tyrphostins also decreased the cell numbers to below control numbers in cultures maintained in 1% BSA or in serum containing medium (cytostatic/cytotoxic effects). B44 (EGFR-selective tyrphostin), AG1295 (platelet-derived growth factor receptor [PDGFR]-selective tyrphostin), and A1 had no inhibitory effects on cells with or without E2 treatments. However, A1 inhibited cell growth under serum supplementation. Genistein, a phytoestrogen, stimulated the autonomous, E2-induced as well as serum-induced growth of MCF-7 cells. Cell proliferation results derived from the MTT assay were corroborated by both the colony formation assay as well as the Ki-67 assay. CONCLUSIONS: Of the agents tested, only EGFR-selective tyrphostins blocked E2-stimulated tumor cell proliferation, as opposed to the PDGFR-selective tyrphostin, RTK noninhibitory agent, or the phytoestrogen, genistein, which did not exert such an effect. These findings suggest that epidermal growth factor (EGF) is an important mediator of E2-induced proliferation of MCF-7 cells. Thus, tyrphostins may be selectively used to prevent the growth of hormone-dependent breast cancers, particularly regrowth of residual tumor in postmenopausal breast cancer survivors receiving estrogen replacement therapy.

Antineoplastic Agents↗