PubMed Health⌕ Search

Biomedical subjects

D Patel

Publications and source records attributed to D Patel.

At least 37 records · Page 2Linked to original sources
PubMed2002

Functional analysis of bone morphogenetic protein type II receptor mutations underlying primary pulmonary hypertension.

A wide range of mutations in the type II receptor for bone morphogenetic protein (BMPR-II) have been shown to underlie primary pulmonary hypertension. To determine the mechanism of altered BMPR-II function, we employed transient transfection studies in cell lines and primary cultures of pulmonary vascular smooth muscle cells using green fluorescent protein (GFP)-tagged wild-type and mutant BMPR2 constructs and confocal microscopy to localize receptors. Substitution of cysteine residues in the ligand binding or kinase domain prevented trafficking of BMPR-II to the cell surface, and reduced binding of (125)I-BMP4. In addition, transfection of cysteine-substituted BMPR-II markedly reduced basal and BMP4-stimulated transcriptional activity of a BMP/Smad responsive luciferase reporter gene (3GC2wt-Lux), compared with wild-type BMPR-II, suggesting a dominant-negative effect of these mutants on Smad signalling. In contrast, BMPR-II containing non-cysteine substitutions in the kinase domain were localized to the cell membrane, although these also suppressed the activity of 3GC2wt-Lux. Interestingly, BMPR-II mutations within the cytoplasmic tail trafficked to the cell surface, but retained the ability to activate 3GC2wt-Lux. Transfection of mutant, but not wild-type, constructs into a mouse epithelial cell line (NMuMG cells) led to activation of p38(MAPK) and increased serum-induced proliferation compared with the wild-type receptor, which was partly p38(MAPK)-dependent. We conclude that mutations in BMPR-II heterogeneously inhibit BMP/Smad-mediated signalling by diverse molecular mechanisms. However, all mutants studied demonstrate a gain of function involving upregulation of p38(MAPK)-dependent proproliferative pathways.

Animals↗
PubMed2002

Rituximab-based chemotherapy for steroid-refractory autoimmune hemolytic anemia of chronic lymphocytic leukemia.

Autoimmune hemolytic anemia (AIHA) is a well known complication of chronic lymphocytic leukemia (CLL). Steroids are the first line of treatment and there are limited effective treatment options for steroid refractory AIHA of CLL. Rituximab, an active agent against B cell malignancies, has also been noted to be active in certain autoimmune hematologic disorders. We used a combination of rituximab, cyclophosphamide and dexamethasone (RCD) in eight CLL patients with steroid refractory AIHA. Rituximab was given at a dose of 375 mg/m(2) i.v. on day 1 (D-1). Cyclophosphamide was given at a dose of 750 mg/m(2) on D-2. Twelve mg of dexamethasone was given i.v. on D-1, D-2 and orally from D-3 to D-7. Cycles were repeated every 4 weeks till the best response. Response in AIHA was evaluated by frequent blood counts and Coombs test. All eight patients achieved a remission of their AIHA. Median pretreatment hemoglobin was 8.3 g/dl and post-treatment hemoglobin was 14.3 g/dl. Five patients converted to Coombs negative after RCD. Median duration of response was 13 months (7-23+). Retreatment with RCD was also effective in achieving a response on relapse of AIHA. Our results indicate that a rituximab-based combination regimen (RCD) is highly effective in treating steroid refractory AIHA of CLL.

Aged↗
PubMed2002

Out of hours management of occupational exposures to blood and body fluids in healthcare staff.

AIMS: To assess and compare the out of hours and in hours management of occupational blood and body fluid exposures in a London teaching hospital. METHODS: The occupational health and accident and emergency records of individuals presenting with occupational body fluid exposures over a six month period at a London teaching hospital were analysed retrospectively. Main outcome measures were the completeness of records, and the appropriate management of body fluid exposures using the Department of Health guidelines as the gold standard. RESULTS: A total of 177 body fluid exposures were reported; 109 (61.58%) were initially assessed in the occupational health department, and 68 (38.42%) in the accident and emergency department. Of those originally assessed in the accident and emergency department, only 21 (30.88%) attended the occupational health department for follow up. Occupational health staff were more consistent in assessing and managing exposures, and in a higher proportion of cases gave more appropriate advice on post-exposure prophylaxis (PEP) against hepatitis B and HIV. Of the 11 individuals prescribed HIV PEP (all by accident and emergency staff), only three subsequently attended occupational health for follow up. In all three cases therapy was discontinued, as the source was HIV negative or the exposure low risk. CONCLUSIONS: Out of hours management of occupational body fluid exposures, particularly the prescribing of HIV PEP, was inconsistent with in hours practice. This may also be the case in other large inner city hospitals offering a similar service.

Blood↗
PubMed2002

Detection of serum M2 anti-mitochondrial antibodies by enzyme-linked immunosorbent assay is potentially less specific than by immunofluorescence.

AIM: To compare the predictive values of enzyme-linked immunosorbent assays (ELISAs) and indirect immunofluorescence (IIF) techniques for the detection of M2 anti-mitochondrial antibodies. METHODS: Commercial ELISAs are widely available for the detection of anti-mitochondrial antibody subtypes in primary biliary cirrhosis (PBC). We compared the results from two ELISAs (one recombinant, one purified antigen) with those from two IIF methods in a well-defined cohort of PBC patients and in patients with systemic lupus erythematosus, Sjögren's syndrome, sicca syndrome, systemic sclerosis, rheumatoid arthritis and blood donor controls. RESULTS: There was good correlation between a rodent substrate IIF and ELISA A (r=0.9134), but poor correlation with ELISA B (r=0.5999), which produced many false-positive results in the control population. We show that rodent IIF alone or human epithelial cell (HEp-2000) screening with confirmation by ELISA produce similar predictive values for PBC and require lesser degrees of skilled interpretation of IIF patterns. CONCLUSIONS: We conclude that the specificities of IIF are greater than the ELISA methods (99% versus 85-97%), although the ELISAs are slightly more sensitive in biopsy-proven PBC. Careful in-house validation of all new ELISA technologies is mandatory for good laboratory practice, but IIF in experienced hands remains an effective and specific assay.

Animals↗
PubMed2001

Connexin26 in adult rodent central nervous system: demonstration at astrocytic gap junctions and colocalization with connexin30 and connexin43.

The connexin family of proteins (Cx) that form intercellular gap junctions in vertebrates is well represented in the mammalian central nervous system. Among these, Cx30 and Cx43 are present in gap junctions of astrocytes. Cx32 is expressed by oligodendrocytes and is present in heterologous gap junctions between oligodendrocytes and astrocytes as well as at autologous gap junctions between successive myelin layers. Cx36 mRNA has been identified in neurons, and Cx36 protein has been localized at ultrastructurally defined interneuronal gap junctions. Cx26 is also expressed in the CNS, primarily in the leptomeningeal linings, but is also reported in astrocytes and in neurons of developing brain and spinal cord. To establish further the regional, cellular, and subcellular localization of Cx26 in neural tissue, we investigated this connexin in adult mouse brain and in rat brain and spinal cord using biochemical and immunocytochemical methods. Northern blotting, western blotting, and immunofluorescence studies indicated widespread and heterogeneous Cx26 expression in numerous subcortical areas of both species. By confocal microscopy, Cx26 was colocalized with both Cx30 and Cx43 in leptomeninges as well as along blood vessels in cortical and subcortical structures. It was also localized at the surface of oligodendrocyte cell bodies, where it was coassociated with Cx32. Freeze-fracture replica immunogold labeling (FRIL) demonstrated Cx26 in most gap junctions between cells of the pia mater by postnatal day 4. By postnatal day 18 and thereafter, Cx26 was present at gap junctions between astrocytes and in the astrocyte side of most gap junctions between astrocytes and oligodendrocytes. In perinatal spinal cord and in five regions of adult brain and spinal cord examined by FRIL, no evidence was obtained for the presence of Cx26 in neuronal gap junctions. In addition to its established localization in leptomeningeal gap junctions, these results identify Cx26 as a third connexin (together with Cx30 and Cx43) within astrocytic gap junctions and suggest a further level of complexity to the heterotypic connexin channel combinations formed at these junctions.

Aging↗
PubMed2001

Insulin-like growth factor-1 induces survival and growth of biologically early melanoma cells through both the mitogen-activated protein kinase and beta-catenin pathways.

Melanoma cells produce growth factors for autocrine growth control and for stimulating fibroblasts and endothelial cells in the tumor stroma. Activated stromal fibroblasts can in turn secrete growth factors that support tumor growth. We studied this feedback from fibroblasts to melanoma cells by overexpressing insulin-like growth factor 1 (IGF-1) with an adenoviral vector. Melanoma cells do not produce IGF-1. IGF-1 enhanced survival, migration, and growth of cells from biologically early lesions, but not from biologically late primary or metastatic lesions. Early melanoma cells were activated by IGF-1 to phosphorylate Erk1 and -2 of the mitogen-activated protein kinase pathway. IGF-1 also activated Akt, inhibited its down-stream effector GSK3-beta, and stabilized beta-catenin. Late primary and metastatic melanoma cells did not respond to growth stimulation by IGF-1 because of a constitutive activation of the mitogen-activated protein kinase pathway and a higher level of stabilized beta-catenin. These studies demonstrate that fibroblast-derived growth factors from the tumor environment can provide the malignant cells with a positive feedback through multiple mechanisms but that this stimulation is required only for cells from early and not late stages of tumor progression.

Cell Division↗
PubMed2001

Mel-CAM-specific genetic suppressor elements inhibit melanoma growth and invasion through loss of gap junctional communication.

Normal human melanocytes are interspersed singly among keratinocytes along the basement membrane of the epidermis, whereas melanoma cells readily adhere to each other during invasion of the dermis or distant organs. The tumorigenic and metastatic phenotype of melanoma cells often correlates with increased expression of cell-cell and cell-matrix adhesion receptors. Mel-CAM (MCAM, MUC 18, CD146) is a cell-cell adhesion receptor highly expressed by melanoma cells but not normal melanocytes. We show here that inhibition of Mel-CAM expression in metastatic melanoma cells using genetic suppressor elements of Mel-CAM cDNA leads to inhibition of adhesion between melanoma cells and to downregulation of the tumorigenic phenotype. Growth was not inhibited in genetic suppressor elements-transduced melanoma cells cultured in monolayers but was inhibited when cells were maintained anchorage-independently in soft agar and greatly reduced in immunodeficient mice. A three-dimensional epidermal skin equivalent model demonstrated that Mel-CAM allows melanoma cells to separate from the epidermis and invade the basement membrane zone and dermis. However, melanoma cells with little or no Mel-CAM were poorly invasive, possibly due to their loss of gap junctional communication. These results suggest the multifunctional role of a melanoma-associated cell-cell adhesion receptor in tumor progression.

Animals↗
PubMed2001

Continuous affinity-based selection: rapid screening and simultaneous amplification of bacterial surface-display libraries.

A new method for continuous biopanning has been developed. We have combined the power of affinity chromatography with the fecundity of bacteria in a unique process that mimics clonal selection. Mixed populations of bacteria were applied to a fermenter containing the immobilized ligand of interest. Bacteria retained in this affinity fermenter were allowed to grow under continuous washout conditions, such that weakly bound organisms were selectively lost. Those initially rare founder bacteria expressing a receptor for the immobilized ligand (R+ve) were thus enriched and amplified simultaneously. From an initial culture containing 1 x 10(10) R-ve cells spiked with fewer than 30 R+ve bacteria (<1 in 10(8)), final ratios of R+ve/R-ve bacteria as high as 1 in 12 were observed, representing an enrichment factor of 55 million-fold. This technology has considerable potential for rapid screening of bacterial surface-display libraries and in facilitating directed-evolution studies.

Bacterial Outer Membrane Proteins↗
PubMed2001

Unusually wide co-factor tolerance in a metalloenzyme; divalent metal ions modulate endo-exonuclease activity in T5 exonuclease.

T5 5'-3' exonuclease is a member of a homologous group of 5' nucleases which require divalent metal co-factors. Structural and biochemical studies suggest that single-stranded DNA substrates thread through a helical arch or hole in the protein, thus bringing the phosphodiester backbone into close proximity with the active site metal co-factors. In addition to the expected use of Mg(2+), Mn(2+) and Co(2+) as co-factors, we found that divalent zinc, iron, nickel and copper ions also supported catalysis. Such a range of co-factor utilisation is unusual in a single enzyme. Some co-factors such as Mn(2+) stimulated the cleavage of double-stranded closed-circular plasmid DNA. Such endonucleolytic cleavage of circular double-stranded DNA cannot be readily explained by the threading model proposed for the cleavage of substrates with free 5'-ends as the hole observed in the crystal structure of T5 exonuclease is too small to permit the passage of double-stranded DNA. We suggest that such a substrate may gain access to the active site of the enzyme by a process which does not involve threading.

Base Sequence↗
PubMed2001

High prevalence of myocardial perfusion abnormalities on positron emission tomography in asymptomatic persons with a parent or sibling with coronary artery disease.

BACKGROUND: We hypothesized that asymptomatic persons with a parent or sibling with coronary artery disease (CAD) have myocardial perfusion defects on positron emission tomography (PET) as markers of early CAD. METHODS AND RESULTS: After medical and family histories were recorded, 90 subjects underwent rest-dipyridamole cardiac PET perfusion imaging, including 18 index cases (a subject with CAD documented by PET and arteriography), 32 asymptomatic adults without known CAD who had a parent or sibling with CAD among these index cases, 30 asymptomatic subjects with comparable coronary risk factors without CAD or a family history of CAD, and 10 volunteer control subjects with no risk factors and no family history. PET perfusion images were quantified with automated software for size of abnormalities as percent of the cardiac image outside 95% CIs of normal controls and for severity as the lowest quadrant average relative activity. Of asymptomatic subjects with a parent or sibling with CAD (first-degree relatives), 50% had dipyridamole-induced myocardial perfusion defects that involved >/=5% of the cardiac image outside normal 95% CIs with or without other risk factors. The size of perfusion defects was larger in first-degree relatives than in control subjects (11+/-13% versus 1+/-1%, P:=0.02) and larger than in asymptomatic subjects with comparable risk factors but no family history of CAD (11+/-13% versus 5+/-6%, P:=0.02). CONCLUSIONS: This study documents the presence of quantitative, statistically significant, dipyridamole-induced myocardial perfusion abnormalities on PET in 50% of asymptomatic persons with a parent or sibling with CAD, independent of other risk factors, indicating preclinical coronary atherosclerosis.

Adult↗
PubMed2001

Intravenous gammglobulin-associated acute renal failure.

Intravenous gammaglobulins are used for the treatment of various auto-immune hematological disorders. Renal failure is a relatively rare, but an increasingly recognized side effect of gammaglobulin therapy. Although the renal failure is usually reversible, renal replacement therapy is required occasionally. A high index of suspicion, early recognition and appropriate intervention can prevent this complication. We herewith describe two patients with an immune hematological disorder, who developed acute renal failure after treatment with intravenous gammaglobulins. A brief review of the possible risk factors, pathophysiology and management of this complication is provided.

Acute Kidney Injury↗
PubMed2001

Spontaneous coronary artery dissection in a pregnant woman.

BACKGROUND: Spontaneous coronary dissection is a rare condition occurring more often in women, with a higher frequency during the peripartum period. No specific etiology has been defined to describe this uncommon, yet often fatal phenomenon. CASE: A young woman presented at 36 weeks of a noncomplicated pregnancy with recent onset of diaphoresis, dyspnea, and tingling substernal chest discomfort. Upon evaluation, she developed cardiovascular collapse and ventricular fibrillation requiring aggressive resuscitative measures, eventually leading to extracorporeal membrane oxygenation. Right coronary artery dissection was ultimately diagnosed and treated with intracoronary angioplasty and stent placement. CONCLUSION: Spontaneous coronary dissection must be considered when evaluating a patient with a similar clinical presentation, given its overall mortality of more than 50% at presentation, particularly in the peripartum period.

Adult↗
PubMed2001

Advances in surgical dentistry.

The introduction of the specialty of surgical dentistry is an advance in itself and has far-reaching implications for the future. Surgical specialists will have a significant impact on the provision and resources of surgical dental services. This paper explores recent developments within the field.

Adult↗
PubMed2001

Effect of a statewide neonatal resuscitation training program on Apgar scores among high-risk neonates in Illinois.

OBJECTIVE: The national Neonatal Resuscitation Program (NRP), started in 1987, provided training to hospital delivery room personnel to standardize knowledge and skills to reduce neonatal morbidity and mortality and increase successful resuscitation during the first few critical minutes after birth. The Apgar score continues to be used as the best established index of immediate postnatal health. The purpose of this study was to evaluate the impact of the NRP instruction in Illinois hospitals by examining Apgar scores among high-risk infants who are likely to benefit from the NRP. METHODS: A retrospective 3-time period cohort design was used (before the introduction of the NRP, 1985-1988; transition when NRP training occurred, 1989-1990; and after NRP training was completed at least once for some delivery room personnel in each Illinois hospital, 1991-1995). Illinois computerized birth certificate files on a selected group of 636 429 high-risk neonates provided information on Apgar scores and maternal characteristics. The American Academy of Pediatrics provided instructor lists to determine when NRP training started and when it was fully implemented in Illinois. Illinois Department of Public Health provided data to categorize hospitals into levels based on type and intensity of neonatal services (Level I, II, II+, III). High-risk neonates were defined as meeting 1 of the following criteria: maternal age <20 years old or >35 years old, birth weight <2500 g or >4000 g, presence of a maternal medical risk factor, and no prenatal care or prenatal care started after the first trimester. Several exclusion criteria were applied including the following: birth records with missing data, multiple birth or congenital anomaly, and hospital information that indicate no birth deliveries in 1 of the 11 study years or delivery outside of a hospital. One-minute and 5-minute Apgar scores were divided into categories for analysis (0-3, 4-6, 7-10). No change or a decrease in a low (0-6) 1-minute Apgar when compared with the 5-minute Apgar was a primary measure to evaluate effect of NRP resuscitation. Variables examined included the following: race/ethnicity, maternal age, level of education, presence of maternal medical risk factor, trimester started prenatal care, complications of labor and delivery, and a low birth weight. Analysis consisted of chi(2) tests, relative risk calculations, and logistic regression to reveal independent associations with no change in low 1-minute Apgar score or continued low (0-6) 5-minute Apgar. RESULTS: A total of 636 429 high-risk birth records was selected for detailed analyses out of 2 077 533 births in Illinois between 1985 and 1995 for 193 hospitals. The number of active NRP instructors in Illinois changed dramatically during the study period; for example, 1 to 6 between 1987 and 1988 to 1096 to 1242 between 1991 and 1995. The percentage of neonates reported to have low (<7) 1-minute Apgar score decreased in 1991 to 1995 overall and for each of 4 hospital levels. Overall and by hospital level, there was a statistically significant lower proportion of high-risk newborns who showed a decrease or no change in their 5-minute Apgar scores after the NRP instruction. After adjusting for several maternal characteristics, logistic regression analysis revealed that high-risk newborns with a low 1-minute Apgar were more likely to increase their 5-minute Apgar after the NRP instruction in 1991 to 1995. Additional analyses indicated that very low birth weight and low birth weight newborns benefited the most from NRP instruction. CONCLUSION: Although previous research has shown that the NRP instruction improves knowledge and skill among health care personnel in the delivery room, both short-term and long-term, there has been little evidence to demonstrate NRP impact on infant morbidity. Several strategies were used in this study to control for bias and to adjust for secular trends in decreased infant morbidity during the study period. This study demonstrated sufficient support for the hypothesis that a significant improvement occurred among neonates in their Apgar score after the NRP instruction in Illinois. Empirical support is provided for the clinical effectiveness of NRP instruction.

Apgar Score↗
PubMed2001

In vitro assay for site-specific proteases using bead-attached GFP substrate.

Site-specific proteases, which catalyze cleavage of peptide bonds in specific amino acid sequences of target proteins, play important roles in various biological events of many living organisms. In humans, disruption in regulation of these site-specific proteases can lead to pathological consequences. Here, we report a simple in vitro assay for enzymatic activities of site-specific proteases. This assay system employs a protein substrate molecule that is comprised of (i) His-tag binding module, (ii) cleavage sites, and (iii) green fluorescent protein (GFP) detection module. In this study, prostate-specific antigen (PSA) and Thrombin-specific cleavage sites were introduced into the substrate molecules. The overexpressed GFP substrate protein was purified with the aid of Ni++-charged magnetic beads. On cleavage by either PSA or Thrombin, GFP was released from the bound magnetic beads, enabling a direct measurement of the cleaved product by fluorescence. Detection sensitivity, as well as the kinetics of reaction of PSA cleavage with the GFP substrate, was similar or better than commercially available PSA fluorogenic peptide substrate. This bead-attached GFP substrate was also used for an inhibition assay using a competitive inhibitor of Thrombin. In conclusion, this assay offers a simple fluorescent method for monitoring the activity of the site-specific proteases. Furthermore, this system provides flexible means of incorporating varying sizes of flanking sequences adjacent to the cleavage site, which can be essential for studying the regulatory macromolecular interactions between proteases and their substrates.

Animals↗
PubMed2001

Protection against drug- and chemical-induced multiorgan toxicity by a novel IH636 grape seed proanthocyanidin extract.

Grape seed proanthocyanidins have been demonstrated to exhibit a broad spectrum of pharmacological, therapeutic and chemoprotective properties. In our previous studies, IH636 grape seed proanthocyanidin extract (GSPE, commercially known as ActiVin) demonstrated excellent concentration- and dose-dependent free radical scavenging abilities in both in vitro and in vivo models and provided significantly better protection than vitamins C, E and beta-carotene. GSPE demonstrated significant cytotoxicity towards human breast, lung and gastric adenocarcinoma cells, while enhancing the growth and viability of normal human gastric mucosal cells and macrophage J774A.1 cells. In this study, the bioavailability and protective ability of GSPE were examined against acetaminophen-induced hepatoxicity, amiodarone-induced pulmonary toxicity, doxorubicin-induced cardiotoxicity, cadmium chloride-induced nephrotoxicity, dimethylnitrosamine-induced spleenotoxicity and O-ethyl-S,S-dipropyl phosphorodithioate (MOCAP)-induced neurotoxicity in mice. In each experiment, half of the test animals were orally fed GSPE for 7-10 days prior to drug/chemical exposure, while the other half received no GSPE. Parameters of analysis included changes in serum chemistry [alanine amino-transferase (ALT), blood urea nitrogen and creatine kinase], histopathology and integrity of genomic DNA. The results indicated that GSPE preexposure prior to the drugs/chemicals such as acetaminophen, amiodarone, doxorubicin, cadmium chloride or dimethylnitrosamine treatment, provided near complete protection in terms of serum chemistry changes (ALT, blood urea nitrogen and creatine kinase) and inhibition of both forms of cell death, e.g., apoptosis and necrosis. DNA damage in various tissues triggered by these agents was significantly reduced. Histopathological examination of the organs evaluated reflected similar patterns to those of the serum chemistry and DNA results. MOCAP exposure showed symptoms of severe neurotoxicity coupled with serum chemistry changes in the absence of any significant genomic change or brain pathology. GSPE afforded only partial protection in the brain tissue. These results suggest that GSPE exposure is bioavailable and provides significant multiorgan protection against drug- and chemical-induced toxic assaults.

Alanine Transaminase↗