Search PubMed⌕ Search

Biomedical subjects

S Gross

Publications and source records attributed to S Gross.

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

Vectors for the expression of tagged proteins in Drosophila.

Regulated expression systems have been extremely useful in developmental studies, allowing the expression of specific proteins in defined spatial and temporal patterns. If these proteins are fused to an appropriate molecular tag, then they can be purified or visualized without the need to raise specific antibodies. If the tag is inherently fluorescent, then the proteins can even be visualized directly, in living tissue. We have constructed a series of P element-based transformation vectors for the most widely used expression system in Drosophila, GAL4/UAS. These vectors provide a series of useful tags for antibody detection, protein purification, and/or direct visualization, together with a convenient multiple cloning site into which the cDNA of interest can be inserted.

Amino Acid Sequence↗
PubMed2001

Differential expression of matrix metalloproteinases and the tissue inhibitor in human milk.

There are differences in the temporal expression of MMP-2 and TIMP-4 in human milk from Day 1 to 30 postpartum in healthy mothers of term pregnancies: (i) MMP-2 activity peaked at Day 1 (colostrum) then exponentially decreased afterwards; and (ii) The expression of TIMP-4 was maximal at Day 7 and persisted thereafter. From our preliminary findings, the differential expression of MMP and the tissue inhibitor in human milk may be related to the protective action of human milk.

Adult↗
PubMed2000

Genotypic and phenotypic spectrum in tricho-rhino-phalangeal syndrome types I and III.

Tricho-rhino-phalangeal syndrome (TRPS) is characterized by craniofacial and skeletal abnormalities. Three subtypes have been described: TRPS I, caused by mutations in the TRPS1 gene on chromosome 8; TRPS II, a microdeletion syndrome affecting the TRPS1 and EXT1 genes; and TRPS III, a form with severe brachydactyly, due to short metacarpals, and severe short stature, but without exostoses. To investigate whether TRPS III is caused by TRPS1 mutations and to establish a genotype-phenotype correlation in TRPS, we performed extensive mutation analysis and evaluated the height and degree of brachydactyly in patients with TRPS I or TRPS III. We found 35 different mutations in 44 of 51 unrelated patients. The detection rate (86%) indicates that TRPS1 is the major locus for TRPS I and TRPS III. We did not find any mutation in the parents of sporadic patients or in apparently healthy relatives of familial patients, indicating complete penetrance of TRPS1 mutations. Evaluation of skeletal abnormalities of patients with TRPS1 mutations revealed a wide clinical spectrum. The phenotype was variable in unrelated, age- and sex-matched patients with identical mutations, as well as in families. Four of the five missense mutations alter the GATA DNA-binding zinc finger, and six of the seven unrelated patients with these mutations may be classified as having TRPS III. Our data indicate that TRPS III is at the severe end of the TRPS spectrum and that it is most often caused by a specific class of mutations in the TRPS1 gene.

Adolescent↗
PubMed2000

Differential inhibition of renal cancer cell invasion mediated by fibronectin, collagen IV and laminin.

Invasion of tumor cells into the extracellular matrix is an essential step in the formation of metastases in renal cancer. Cell adhesion molecules such as beta(1)-integrins, which bind to the RGD sequence (arginine-glycine-asparagine) and CD44 are involved in this process. We examined the invasion of a renal carcinoma cell line (CCF-RC1) into the extracellular matrix compounds fibronectin, collagen IV and laminin and the effect of TGFbeta and IFNgamma on this process. The inhibitory effect of an antibody against the beta(1)-subunit of integrins (CD29), as well as a pentapeptide including the RGD sequence, was also evaluated. A micro-chemotaxis chamber, including a polycarbonate membrane with a pore diameter of 8 microm, was used for quantification of cell migration. The addition of the extracellular matrix compounds fibronectin, laminin and collagen IV resulted in a 5-10-fold increase in invasion. This increased invasion depends strongly on the presence of beta(1)-integrins, shown by the use of an antibody against CD29 or a RGD including peptide which inhibit the cell migration by approximately 88%. CD44 is less involved in collagen IV dependent migration and almost no influence of CD44 was observed on a fibronectin and laminin dependent migration. TNFalpha and IFNgamma did not significantly influence the expression of CD29 or CD44, and no alteration in tumor cell migration was observed. These results show that the invasion of renal cancer cells is differentially regulated by compounds of the extracellular matrix, whereby fibronectin seems to be the most critical factor. The molecular interactions in this process are strongly dependent on beta(1)-integrins and the corresponding amino acid sequence RGD.

Carcinoma, Renal Cell↗
PubMed2000

Site-selective dephosphorylation of the platelet-derived growth factor beta-receptor by the receptor-like protein-tyrosine phosphatase DEP-1.

Ligand stimulation of PDGF beta-receptors leads to autophosphorylation of the regulatory tyrosine 857 and of tyrosine residues that in their phosphorylated form serve as docking sites for Src homology 2 domain-containing proteins. Regulation of the PDGF beta-receptor by protein-tyrosine phosphatases is poorly understood. We have investigated PDGF beta-receptor dephosphorylation by receptor-like protein-tyrosine phosphatase DEP-1 using a cell line with inducible DEP-1 expression and by characterizing in vitro dephosphorylation of the PDGF beta-receptor and of receptor-derived phosphopeptides by DEP-1. After DEP-1 induction PDGF beta-receptor.DEP-1 complexes and reduced receptor tyrosine phosphorylation were observed. Phosphopeptide analysis of the PDGF beta-receptors from DEP-1-expressing cells and of the receptors dephosphorylated in vitro by DEP-1 demonstrated that dephosphorylation of autophosphorylation sites of the receptor differed and revealed that the regulatory Tyr(P)(857) was not a preferred site for DEP-1 dephosphorylation. When dephosphorylation of synthetic receptor-derived peptides was analyzed, the selectivity was reproduced, indicating that amino acid sequence surrounding the phosphorylation sites is the major determinant of selectivity. This notion is supported by the observation that the poorly dephosphorylated Tyr(P)(562) and Tyr(P)(857), in contrast to other analyzed phosphorylation sites, are surrounded by basic amino acid residues at positions -4 and +3 relative to the tyrosine residue. Our study demonstrates that DEP-1 dephosphorylation of the PDGF beta-receptor is site-selective and may lead to modulation, rather than general attenuation, of signaling.

Amino Acid Sequence↗
PubMed2000

Identification of a testis-specific gene (C15orf2) in the Prader-Willi syndrome region on chromosome 15.

Prader-Willi syndrome (PWS) results from the loss of paternal contributions for a 2-Mb imprinted region on the proximal long arm of human chromosome 15. Hitherto, five paternally active genes have been identified in this region (ZNF127, NDN, MAGEL2, SNURF-SNRPN, and IPW). Here we report the identification of a novel gene in the PWS critical region, which has been designated "chromosome 15 open reading frame 2" (C15orf2). C15orf2 is an intronless gene located between MAGEL2 and SNURF-SNRPN. It is associated with a CpG island, which is methylated in all tissues tested except for germ cells. C15orf2 is transcribed as a 7.5-kb mRNA and contains an open reading frame encoding a predicted 1156-amino-acid protein of unknown function. Transcription of C15orf2 occurs exclusively in the testis, and in adult testis samples, we observed biallelic expression. By zoo-blot analysis, we found related sequences in DNA from other primates, but not in nonprimate DNA. We conclude that C15orf2 may play a role in primate spermatogenesis.

3' Untranslated Regions↗
PubMed2000

Protein phosphatase 1beta is required for the maintenance of muscle attachments.

Type 1 serine/threonine protein phosphatases (PP1) are important regulators of many cellular and developmental processes, including glycogen metabolism, muscle contraction, and the cell cycle [1] [2] [3] [4] [5]. Drosophila and humans both have multiple genes encoding PP1 isoforms [3] [6] [7]; each has one beta and several alpha isoform genes (alpha(1), alpha(2), alpha(3) in flies, alpha and gamma in humans; mammalian PP1beta is also known as PP1delta). The alpha/beta subtype differences are highly conserved between flies and mammals [6]. Though all these proteins are >85% identical to each other and have indistinguishable activities in vitro, we show here that the Drosophila beta isoform has a distinct biological role. We show that PP1beta9C corresponds to flapwing (flw), previously identified mutants of which are viable but flightless because of defects in indirect flight muscles (IFMs) [8]. We have isolated a new, semi-lethal flw allele that shows a range of defects, especially in muscles, which break away from their attachment sites and degenerate.

Alleles↗
PubMed2000

Electrospray ionization in the study of the polycondensation of Ti(O-i-C3H7)4 and Ti(O-n-C4H9)4

The polycondensation of Ti(O-i-C3H7)4 (1) and Ti(O-n-C4H9)4 (2), precursors widely employed in sol-gel processes, has been investigated by electrospray ionization mass spectrometry. By analysis of 10(-6) M methanol solutions of compounds 1 and 2, the same ionic species are detected, proving that the first step in the polycondensation reaction is the i-propyl (or n-butyl) alcohol-methanol complete exchange. This reaction leads to the Ti(OCH3)4 (3) species, representing the synthon of the polycondensation. Various oligomers of 3 have been detected and characterized by MS/MS experiments, and the related mechanisms have been discussed. A minor oligomeric series due to hydroxyl-containing polycondensation products has also been characterized.

Journal Article↗
PubMed2000

Increased expansion and differentiation of cord blood products using a two-step expansion culture.

OBJECTIVE: [corrected] The use of allogeneic cord blood (CB) products as a source of cellular support for patients receiving high-dose chemotherapy has been limited primarily to smaller children due to the low numbers of cells in a CB unit. Ex vivo expansion of CB cells has been proposed as a method to increase the number of cells available for transplantation. Following high-dose chemotherapy administration, we transplanted adult patients with CB expanded in static culture for 10 days, in DM containing stem cell factor (SCF), granulocyte colony-stimulating factor (G-CSF), and megakaryocyte growth and development factor (MGDF). Patients achieved neutrophil engraftment in a median of 26 days (range 15 to 45). In an attempt to hasten the time to neutrophil engraftment, we developed a two-step culture system that results in increased expansion of total nucleated cells and further maturation of neutrophil precursors. MATERIALS AND METHODS: CD34(+) cells isolated from CB products were cultured for 7 days at 37 degrees C in 100-mL Teflon culture bags containing 50 mL of DM containing SCF, G-CSF, and MGDF (100 ng/mL). The cells were harvested from these bags after 7 days of incubation at 37 degrees C and transferred to 1-L Teflon bags containing 1 L of DM plus SCF, G-CSF, and MGDF. After a second culture period of 7 days, the cells were harvested, washed, and assayed for mature (granulocyte-macrophage colony-forming cells [GM-CFC]) and primitive progenitor cells (high proliferative potential colony-forming cells [HPP-CFC]). RESULTS: The two-step cultures resulted in a median total nucleated cell expansion of 438-fold (range 286 to 952, N = 11); the original one-step cultures resulted in a median expansion of 98-fold (range 59 to 350, N = 5). Equivalent expansion of committed progenitor cells (GM-CFC) and primitive progenitor cells (HPP-CFC) was obtained. CD34(+) cells were expanded a median of 29-fold in the two-step cultures (N = 11). The two-step culture contained more mature neutrophil cells, by morphologic examination, than the one-step cultures, similar to ex vivo expanded peripheral blood progenitor cells (PBPC). CONCLUSION: The two-step ex vivo expansion conditions described for CB resulted in increased numbers of total nucleated cells, GM-CFC, HPP-CFC, and CD34(+) cells and morphologically resembled ex vivo expanded PBPC, which have been shown to provide more rapid neutrophil engraftment than unexpanded PBPC. We propose that the availability of increased numbers of expanded CB cells may result in more rapid engraftment of neutrophils following infusion to transplant recipients.

Antigens, CD34↗
PubMed2000

2H-Thieno[3,2-e]- and [2,3-e]-1,2-thiazine-6-sulfonamide 1,1-dioxides as ocular hypotensive agents: synthesis, carbonic anhydrase inhibition and evaluation in the rabbit.

Novel non-chiral 2H-thieno[3,2-e]- and [2,3-e]-1,2-thiazine-6-sulfonamide 1,1-dioxides were synthesized for evaluation as potential candidates for the treatment of glaucoma. All of the compounds prepared were potent high affinity inhibitors of human carbonic anhydrase II, Ki < 0.5 nM. Additionally, inhibition of recombinant human carbonic anhydrase IV was determined for selected compounds; these were shown to be moderate to potent inhibitors of this isozyme with IC50 values ranging from 4.25 to 73.6 nM. Of the compounds evaluated for their ability to lower intraocular pressure in naturally hypertensive Dutch-belted rabbits, 5a, 17a3, 17b1, 17b2, 17h2 and 17i1 showed significant efficacy (> 20% decrease) in this model following topical ocular administration.

Animals↗
PubMed2000

Na+/K+-ATPase inhibition during cardiac myocyte swelling: involvement of intracellular pH and Ca2+.

Previous studies in chick embryo cardiac myocytes have shown that the inhibition of Na+/K+-ATPase with ouabain induces cell shrinkage in an isosmotic environment (290 mOsm). The same inhibition produces an enhanced RVD (regulatory volume decrease) in hyposmotic conditions (100 mOsm). It is also known that submitting chick embryo cardiomyocytes to a hyperosmotic solution induces shrinkage and a concurrent intracellular alkalization. The objective of this study was to evaluate the involvement of intracellular pH (pHi), intracellular Ca2+ ([Ca2+]i) and Na+/K+-ATPase inhibition during hyposmotic swelling. Changes in intracellular pH and Ca2+ were monitored using BCECF and fura-2, respectively. The addition of ouabain (100 microM) under both isosmotic and hyposmotic stimuli resulted in a large increase in [Ca2+]i (200%). A decrease in pHi (from 7.3 +/- 0.09 to 6.4 +/- 0.08, n = 6; p < 0.05) was only observed when ouabain was applied during hyposmotic swelling. This acidification was prevented by the removal of extracellular Ca2+. Inhibition of Na+/H+ exchange with amiloride (1 mM) had no effect on the ouabain-induced acidification. Preventing the mitochondrial accumulation of Ca2+ using CCCP (10 microM) resulted in a blockade of the progressive acidification normally induced by ouabain. The inhibition of mitochondrial membrane K+/H+ exchange with DCCD (1 mM) also completely prevented the acidification. Our results suggest that intracellular acidification upon cell swelling is mediated by an initial Ca2+ influx via Na+/Ca2+ exchange, which under hyposmotic conditions activates the K+ and Ca2+ mitochondrial exchange systems (K+/H+ and Ca2+/H+).

Amiloride↗
PubMed2000

Hierarchical dependency models for multivariate survival data with censoring.

A family of partial likelihood logistic models is proposed for clustered survival data that are reported in discrete time and that may be censored. The possible dependence of individual survival times within clusters is modeled, while distinct clusters are assumed to be independent. Two types of clusters are considered. First, all clusters have the same size and are identically distributed. Second, the clusters may vary in size. In both cases our asymptotic results apply to a large number of small independent clusters.

Antineoplastic Agents↗
PubMed2000

New York State universal newborn hearing screening demonstration project: inpatient outcome measures.

OBJECTIVE: To evaluate the feasibility of universal newborn hearing screening by examining inpatient outcome measures from 8 hospitals located in geographically diverse areas of New York State over a 3-yr period. DESIGN: Funding was provided by the New York State Department of Health to implement predischarge hearing screening programs in the neonatal intensive care units (NICUs) and well-baby nurseries (WBNs) of eight hospitals. Various screening protocols including transient evoked otoacoustic emissions alone or in combination with conventional auditory brain stem response or screening auditory brain stem response were implemented by each site. Measured outcomes included rate of misses, refusals, and fails. Results were analyzed as a function of year of operation, nursery type, and geographic location. RESULTS: Six out of eight hospitals successfully implemented universal hearing screening during the first year, and the remaining 2 hospitals implemented programs during the second year of the project. Over a period of 3 yr, 69,761 newborns were screened at the eight hospitals representing 96.9% of all live births. The overall fail rate (4.04%) combined with the miss rate (2.61%) resulted in 6.63% of infants referred for outpatient follow-up. Mean data indicated that inpatient outcome measures improved with year of operation, with most individual hospitals also showing improvements. Both fail and miss rates were higher in the NICU than in the WBN and for hospitals located in New York City than in other regions of the state. CONCLUSIONS: Inpatient outcome measures of a universal newborn hearing screening project, which involved multiple centers across geographically diverse regions of New York State, were acceptable in terms of successfully screening a high percentage of live births and attaining low refer rates for outpatient screening. This study adds to the growing body of literature supporting the feasibility of screening all newborns before hospital discharge.

Acoustic Stimulation↗
PubMed2000

The New York State universal newborn hearing screening demonstration project: outpatient outcome measures.

OBJECTIVE: To investigate outpatient outcome measures of a multi-center, state-wide, universal newborn hearing screening project. DESIGN: Eight hospitals participated in a 3-yr, funded project. Each hospital designed its own protocol using common criteria for judging whether an infant passed a hearing screening. Infants were tested in the hospital, and those either failing the in-hospital screening or who were not tested in the hospital (missed) were asked to return 4 to 6 wk after hospital discharge for outpatient rescreening. Those infants failing the outpatient rescreening were referred for diagnostic auditory brain stem response testing. Each hospital used its own audiological equipment and criteria to determine whether a particular infant had a hearing loss. All data were collected and analyzed for individual hospitals, as well as totaled across all hospitals. Data were analyzed in terms of year of program operation, nursery type, and geographic region. RESULTS: Seventy-two percent of infants who failed the in-hospital screening returned for outpatient testing. The percentage of in-hospital fails returning for retesting was significantly higher than the percentage of in-hospital misses returning for retesting. The percentage of infants returning for retesting increased with successive years of program operation. Some differences were noted in the percentage of infants returning for retesting among hospitals and geographic regions of the state. Some differences in outpatient outcome measures also were noted between infants originally born into the neonatal intensive care unit (NICU) and the well-baby nursery (WBN). The percentage of infants from the NICU who returned for retesting was slightly higher than that for infants from the WBN. The percentage of infants from the WBN passing the outpatient rescreening was higher than that for the NICU infants. The overall prevalence of hearing loss was 1.96/1000, with that in the NICU being 8/1000 and that in the WBN being 0.9/1000. Positive predictive value for permanent hearing loss based on inpatient screening was approximately 4% and based on outpatient rescreening was approximately 22%. CONCLUSIONS: Several outpatient outcome measures changed with successive years of program operation, suggesting that programs improve over time. Also, some outpatient outcome measures differ between NICU and WBN populations. The differences noted across regions of the state in the percentage of infants returning for outpatient retesting require further research to determine whether differences are due to demographic and/or procedural differences.

Ambulatory Care↗
PubMed2000

The New York State universal newborn hearing screening demonstration project: ages of hearing loss identification, hearing aid fitting, and enrollment in early intervention.

OBJECTIVE: To determine the ages of hearing loss identification, hearing aid fitting, and enrollment in early intervention through a multi-center, state-wide universal newborn hearing screening project. DESIGN: Universal newborn hearing screening was conducted at eight hospitals across New York State. All infants who did not bilaterally pass hearing screening before discharge were recalled for outpatient retesting. Inpatient screening and outpatient rescreening were done with transient evoked otoacoustic emissions and/or auditory brain stem response testing. Diagnostic testing was performed with age appropriate tests, auditory brain stem response and/or visual reinforcement audiometry. Infants diagnosed with permanent hearing loss were considered for hearing aids and early intervention. Ages of hearing loss identification, hearing aid fitting, and enrollment in early intervention were investigated regarding nursery type, risk status, unilateral versus bilateral hearing loss, loss type, loss severity, and state regions. RESULTS: The prevalence of infants diagnosed with permanent hearing loss was 2.0/1000 (85 of 43,311). Of the 85 infants with hearing loss, 61% were from neonatal intensive care units (NICUs) and 67% were at risk for hearing loss. Of the 36 infants fitted with hearing aids, 58% were from NICUs and 78% were at risk for hearing loss. The median age at identification and enrollment in early intervention was 3 mo. Median age at hearing aid fitting was 7.5 mo. Median ages at identification were less for infants from the well-baby nurseries (WBNs) than for the NICU infants and for infants with severe/profound than for infants with mild/moderate hearing loss, but were similar for not-at-risk and at-risk infants. Median ages at hearing aid fitting were less for well babies than for NICU infants, for not-at-risk infants than for at-risk infants, and for infants with severe/ profound hearing loss than for infants with mild/ moderate hearing loss. However, median ages at early intervention enrollment were similar for nursery types, risk status, and severity of hearing loss. CONCLUSIONS: Early ages of hearing loss identification, hearing aid fitting, and enrollment in early intervention can be achieved for infants from NICUs and WBNs and for infants at risk and not at risk for hearing loss in a large multi-center universal newborn hearing screening program.

Age Factors↗
PubMed2000

New York State universal newborn hearing screening demonstration project: effects of screening protocol on inpatient outcome measures.

OBJECTIVE: To examine differences among various test protocols on the fail rate at hospital discharge for infants in the well-baby nursery (WBN) and neonatal intensive care unit (NICU) who received hearing screening through a universal newborn hearing screening demonstration project. DESIGN: The outcomes of several screening protocols were examined. Two technologies were used: transient evoked otoacoustic emissions (TEOAEs) alone or in combination with the auditory brain stem response (ABR). The performance of test protocols in both nurseries within eight hospitals was examined over a 2- to 3-yr period. In the WBN, six hospitals used a screening protocol of TEOAE technology first followed by an ABR (automated or conventional) technology screening for newborns who referred on TEOAE screening. Two hospitals used TEOAE only in the WBN. Seven hospitals used screening protocols in the NICU that used a combination of TEOAE and ABR technologies (TEOAE technology administered first or second, before or after TEOAE, or TEOAE and ABR tests on all infants). Only one hospital used TEOAE technology exclusively for hearing screening. RESULTS: Significant differences among screening protocols were found across hospitals in the first, second, and third years of the program. The combination of TEOAE technology and ABR technology (a two-technology screening protocol) resulted in a significantly lower fail rate at hospital discharge than the use of a single-technology (TEOAE). Fail rates at discharge were twice as high using the one-technology protocol versus two-technology protocol, even when the best outcomes from program year 3 were considered exclusively. Results of two-technology versus one-technology protocols were similar in the NICU. Use of a second technology for screening TEOAE fails significantly reduced every hospital that used the protocol's fail rate at discharge. CONCLUSIONS: A two-technology screening protocol resulted in significantly lower fail rates at hospital discharge in both the WBN and NICU nurseries than use of a single-technology (TEOAE) hearing screening protocol.

Evoked Potentials, Auditory, Brain Stem↗
PubMed2000

A multicenter long-term safety and efficacy trial of preterm formula supplemented with long-chain polyunsaturated fatty acids.

BACKGROUND: The tissue accretion of long-chain polyunsaturated fatty acids is compromised in infants born prematurely. Human milk contains long-chain polyunsaturated fatty acids, but most preterm infant formulas do not. The long-term effects of preterm formula supplemented with arachidonic acid and docosahexaenoic acid, in proportions typical of those in human milk, were therefore investigated. METHODS: In this double-blind, randomized study, 288 preterm infants received experimental formula (n = 77), unsupplemented (control) formula (n = 78), or human milk (n = 133) until 48 weeks postconceptional age (PCA). Term formula, without supplemental long-chain polyunsaturated fatty acids, was administered from 48 to 92 weeks PCA to formula-fed infants and to infants weaned from human milk. Anthropometric and fatty acid data were assessed by using analysis of variance. RESULTS: At 92 weeks PCA, no statistically significant anthropometric measurement differences were found except for midarm circumference, which was smaller in human milk-fed infants than in those fed formula. Phospholipid concentrations were similar in the experimental and human milk-fed groups, and docosahexaenoic acid levels were significantly greater than in the control group. The types and incidences of adverse events were similar among the feeding groups. CONCLUSIONS: The results of this study demonstrate the efficacy and long-term safety of preterm formula supplemented with long-chain polyunsaturated fatty acids.

Aging↗