A community genetics approach to population screening in India for mental retardation--a model for developing countries.

Some 0.55 million people living in semi-urban and slum populations were screened for mental retardation by trained primary health centre (PHC) doctors, nurses and community health volunteers (CHVs). The staff were provided with prior training on the detection, prevention and diagnosis of mental retardation, prenatal diagnosis, and reproductive responsibilities. Field visits were employed to confirm diagnosed developmental disabilities, and demographic data incorporating social maps of 14 PHCs were prepared. Cases with high-risk genetic factors detected by PHC staff were referred to the Centre for Research in Mental Retardation (CREMERE) for cytogenetic and metabolic investigations, thus linking the study population and the Referral Centre. A genetic team interacted with the patient and family members for genetic counselling. Mental retardation was confirmed in 511 of the 525 cases reported, reflecting the positive impact of training on the CHVs. Potentially preventable environmental factors, such as birth asphyxia, infections, and low birth weight were identified in 251 cases (49%), 137 (27%) of which had additional genetic factors. Genetic causes were found in 186 (36%) individuals, the most common being Down syndrome. The study illustrates the urgent need for the integration of genetic screening into the public health services in India.

Distribution of laminins in the basement membranes of the upper gastrointestinal tract and Barrett's oesophagus.

Barrett's oesophagus predisposes to oesophageal adenocarcinoma. In vitro, laminin, a component of the epithelial basement membrane (BM), is important in regulation of cell differentiation. There is limited information on the distribution of laminin chains in the upper gastrointestinal tract (GIT) and none in Barrett's oesophagus. This study aimed to investigate qualitatively the distribution of laminins in the normal upper GIT mucosa and Barrett's oesophagus in order to understand the role of laminins in metaplasia. Immunoperoxidase staining for laminin chains alpha1, alpha2, alpha3, alpha5, beta1, beta2, beta3, gamma1, and gamma2 was performed on frozen endoscopic squamous and Barrett's oesophageal biopsies and surgical resection specimens from squamous oesophagus (in resection specimens for oesophageal cancer), and in oesophageal and gastric biopsies from control subjects. alpha1 laminin was expressed in the BM of submucosal glands and ducts in squamous oesophagus and Brunner's glands in the duodenum, but not in Barrett's oesophagus or elsewhere in the upper GIT. alpha2 laminin chain was expressed in a granular distribution in the BM of squamous epithelium. In columnar epithelium, including Barrett's oesophagus, alpha2 laminin chain was expressed continuously in the BM of glands and deeper pits, but expression was reduced and granular in the surface epithelial BM. beta2 laminin was continuous in squamous epithelial BM, but in Barrett's and cardia, gastric body, and duodenum, it was expressed faintly in the surface but continuously in the BM of glands and deeper pits. The constituents of laminin-5 were continuously expressed in the BM of squamous epithelium, but in the cardia, gastric body, duodenum, and Barrett's, they were expressed only in the BM of surface epithelium, with a sharp decline in the glandular and deeper pit BM. Site-specific distribution of the alpha2 and beta2 laminin chains may therefore have an important role in Barrett's metaplasia. However, the absence of alpha1 laminin in Barrett's mucosa suggests that this is unlikely to play an important role in columnar metaplasia.

Methylene blue staining: is it really useful in Barrett's esophagus?

BACKGROUND: If areas of specialized intestinal metaplasia (SIM) or dysplasia in Barrett's esophagus can be identified at endoscopy, the number of biopsies could be reduced and the sensitivity of biopsy surveillance would increase. It has been suggested that methylene blue (MB) dye staining may be useful for this purpose. METHODS: Nine patients were prospectively studied with Barrett's esophagus. Staining involved sequential spraying of 10% N-acetylcysteine, 0.5% MB and water. Quadrantic biopsies were obtained from Barrett's epithelium and collected in separate containers depending on whether they were taken from stained or unstained areas. Seven patients undergoing yearly surveillance were asked to compare the discomfort of this endoscopy with that of previous surveillance endoscopies. Biopsies were analyzed for the presence and the percentage of SIM and dysplasia by a nonblinded pathologist. RESULTS: MB staining prolonged endoscopy by a mean of 8 minutes (47% increase in procedure time) and was associated with significant vomiting during the procedure in 2 patients. Staining was observed in all 9 patients. All 7 patients undergoing yearly endoscopic surveillance indicated more discomfort with endoscopy plus MB staining. Of 37 biopsies from stained mucosa, 20 contained SIM; of 23 from unstained mucosa, 15 contained SIM (57% sensitivity, 32% specificity for MB staining). CONCLUSIONS: In this small, nonblinded study MB staining was associated with prolongation of endoscopy, increased patient discomfort, and potentially serious complications and was neither very sensitive nor specific for SIM. It is our recommendation that this technique should not be routinely used in endoscopic surveillance of patients with Barrett's esophagus. Further studies of MB staining are needed.

Inborn errors of metabolism discovered in Asian department of pediatrics and mental retardation research center.

To heighten the effectiveness of chemical diagnosis for inborn errors of metabolism (IEM) using urease pretreatment and GC-MS analysis, a sample collection and transportation method was contrived. The resulting "filter paper set" allows simple urine collection and transportation, and enables anyone from anywhere to receive the GC-MS analysis without the limitations of place or time. Using filter paper sets, high-risk screening of undiagnosed children or mentally retarded children with unknown cause was conducted in cooperation with hospitals and universities in several Asian countries. During 8 months 203 patients from China and India were analyzed and 20 cases of IEM were chemically diagnosed. These diagnoses greatly contributed to the treatment of children with intractable diseases who lived in Asian countries where analytical techniques and facilities for IEM were not sufficient.

Asparagine-proline sequence within membrane-spanning segment of SREBP triggers intramembrane cleavage by site-2 protease.

The NH(2)-terminal domains of membrane-bound sterol regulatory element-binding proteins (SREBPs) are released into the cytosol by regulated intramembrane proteolysis, after which they enter the nucleus to activate genes encoding lipid biosynthetic enzymes. Intramembrane proteolysis is catalyzed by Site-2 protease (S2P), a hydrophobic zinc metalloprotease that cleaves SREBPs at a membrane-embedded leucine-cysteine bond. In the current study, we use domain-swapping methods to localize the residues within the SREBP-2 membrane-spanning segment that are required for cleavage by S2P. The studies reveal a requirement for an asparagine-proline sequence in the middle third of the transmembrane segment. We propose a model in which the asparagine-proline sequence serves as an NH(2)-terminal cap for a portion of the transmembrane alpha-helix of SREBP, allowing the remainder of the alpha-helix to unwind partially to expose the peptide bond for cleavage by S2P.

ER stress induces cleavage of membrane-bound ATF6 by the same proteases that process SREBPs.

ATF6 is a membrane-bound transcription factor that activates genes in the endoplasmic reticulum (ER) stress response. When unfolded proteins accumulate in the ER, ATF6 is cleaved to release its cytoplasmic domain, which enters the nucleus. Here, we show that ATF6 is processed by Site-1 protease (S1P) and Site-2 protease (S2P), the enzymes that process SREBPs in response to cholesterol deprivation. ATF6 processing was blocked completely in cells lacking S2P and partially in cells lacking S1P. ATF6 processing required the RxxL and asparagine/proline motifs, known requirements for S1P and S2P processing, respectively. Cells lacking S2P failed to induce GRP78, an ATF6 target, in response to ER stress. ATF6 processing did not require SCAP, which is essential for SREBP processing. We conclude that S1P and S2P are required for the ER stress response as well as for lipid synthesis.

Effect of elevated temperature and light on the stability of butorphanol tartrate.

The purpose of this investigation was to determine the short-term stability of butorphanol tartrate in presence of diluents. A 10-mg/mL solution of butorphanol tartrate was diluted to 5 mg/mL using normal saline, 5% dextrose in water (D5W), or sterile water for injection. The diluted solutions were divided into two groups. The effect of temperature was tested by placing one group of sealed amber vials at room temperature and at 37 degrees C. The effect of light was studied by placing a second group in amber and clear vials, then exposing them directly to light. At regular time intervals over a period of 5 weeks, the solutions were analyzed for butorphanol tartrate and degradation products using a high-performance liquid chromatography assay. The concentration of butorphanol tartrate remained practically unchanged, indicating that butorphanol tartrate is not affected by heat or light in the presence of any of the diluents over a period of 5 weeks.

Second-site cleavage in sterol regulatory element-binding protein occurs at transmembrane junction as determined by cysteine panning.

In response to sterol deprivation, two sequential proteolytic cleavages release the NH2-terminal fragments of sterol regulatory element-binding proteins (SREBPs) from cell membranes. The fragments translocate to the nucleus where they activate genes involved in cholesterol and fatty acid metabolism. The SREBPs are bound to membranes in a hairpin fashion. The NH2-terminal and COOH-terminal domains face the cytoplasm, separated by two membrane spanning segments and a short lumenal loop. The first cleavage occurs at Site-1 in the lumenal loop. The NH2-terminal fragment is then released by cleavage at Site-2, which is believed to lie within the first transmembrane segment. Here, we use a novel cysteine panning method to identify the second cleavage site (Site-2) in human SREBP-2 as the Leu484-Cys485 bond that lies at the junction between the cytoplasmic NH2-terminal fragment and the first transmembrane segment. We transfected cells with cDNAs encoding fusion proteins with single cysteine residues at positions to the NH2-terminal and COOH-terminal sides of cysteine 485. The NH2-terminal fragments were tested for susceptibility to modification with Nalpha-(3-maleimidylpropionyl)biocytin, which attaches a biotin group to cysteine sulfhydryls. Cysteines to the NH2-terminal side of cysteine 485 were retained on the NH2-terminal fragment, but cysteines to the COOH-terminal side of leucine 484 were lost. Leucine 484 is three residues to the COOH-terminal side of the tetrapeptide Asp-Arg-Ser-Arg, which immediately precedes the first transmembrane segment and is required for Site-2 cleavage.

Wheelchair use: a physical sign in gastroenterological practice.

Diagnosis of functional abdominal pain requires exclusion of organic causes, and many patients undergo considerable investigation. A positive physical sign supporting a functional diagnosis could therefore be of benefit. Wheelchair use specifically for abdominal symptoms was suspected to represent such a sign. Review of 300 consecutive new referrals to a gastroenterology clinic revealed 10 wheelchair users. In four women the chair was used because of the abdominal condition. The final diagnosis (with follow-up to at least 12 months) was functional abdominal pain in each of these cases. All four had had surgery without symptom relief, and all had used their chairs intermittently (mainly for social occasions and hospital visits) for at least 12 months. They believed that normal walking was rendered impossible by abdominal pain whereas the other six wheelchair users gave a clear account of lower limb pain or weakness. Secondary gain with reinforcement of the 'sick role' was felt to be the probable explanation for wheelchair use in the former group. Wheelchair attendance at the gastroenterology clinic, in the absence of lower limb symptoms, is a rare observation but one that may usefully be added to criteria for diagnosis of a functional disorder.

Esophageal dilation in pediatric patients using balloon along the side of the endoscope.

Balloon dilation is an effective modality of treatment for esophageal strictures. As standard through-the-scope balloons do not pass through the biopsy channel of pediatric endoscopes, we have developed a technique by which these balloons can be used in pediatric patients by passing them alongside the endoscope. We report our experience of dilation in four patients using this technique.

Synergistic regulation of human beta-globin gene switching by locus control region elements HS3 and HS4.

Proper tissue- and developmental stage-specific transcriptional control over the five genes of the human beta-globin locus is elicited in part by the locus control region (LCR), but the molecular mechanisms that dictate this determined pattern of gene expression during human development are still controversial. By use of homologous recombination in yeast to generate mutations in the LCR within a yeast artificial chromosome (YAC) bearing the entire human beta-globin gene locus, followed by injection of each of the mutated YACs into murine ova, we addressed the function of LCR hypersensitive site (HS) elements 3 and 4 in human beta-globin gene switching. The experiments revealed a number of unexpected properties that are directly attributable to LCR function. First, deletion of either HS3 or HS4 core elements from an otherwise intact YAC results in catastrophic disruption of globin gene expression at all erythroid developmental stages, despite the presence of all other HS elements in the YAC transgenes. If HS3 is used to replace HS4, gene expression is normal at all developmental stages. Conversely, insertion of the HS4 element in place of HS3 results in significant expression changes at every developmental stage, indicating that individual LCR HS elements play distinct roles in stage-specific beta-type globin gene activation. Although the HS4 duplication leads to alteration in the levels of epsilon- and gamma-globin mRNAs during embryonic erythropoiesis, total beta-type globin mRNA synthesis is balanced, thereby leading to the conclusion that all of the human beta-locus genes are competitively regulated. In summary, the human beta-globin HS elements appear to form a single, synergistic functional entity called the LCR, and HS3 and HS4 appear to be individually indispensable to the integrity of this macromolecular complex.

Rectal lymphogranuloma venereum in association with rectal adenocarcinoma.

Rectal involvement in lymphogranuloma venereum (LGV) is more common in women. Inguinal bubo is often absent and the patient seeks medical attention only at a late stage when rectal stricture has developed. LGV rectal stricture resembles and is known to predispose to rectal cancer. Hence it is necessary to rule out rectal malignancy in patients with LGV stricture. We report a case of rectal LGV associated with rectal adenocarcinoma.

Dilation of esophageal strictures induced by radiation therapy for cancer of the esophagus.

During a 2-year period, 103 consecutive patients undergoing dilation of esophageal strictures induced by radiation therapy for cancer of the esophagus were prospectively studied. The length of the strictures ranged from 0.5 to 13.5 cm (median, 5 cm) and the luminal diameter from 1 to 11 mm (median, 6 mm). Patients were referred for dilation from 2 weeks to 5 years (median, 2 months) after completion of radiation therapy. The guide wire was placed using fluoroscopy in 21 patients, endoscopy in 61, and a combination of endoscopy and fluoroscopy in 21. At least one dilator larger than the stricture could be passed in 101 (98%) patients. Five strictures were dilated to 16 mm, 29 to 15 mm, 28 to 14 mm, 16 to 12.8 mm, and 23 to 12 mm or less during the initial procedure. Development of complications and severe resistance were the limiting factors for optimal dilation. Relief of dysphagia was adequate in 66% of patients. The duration of dysphagia relief was 3 to 84 weeks (median, 16 weeks). Complications included persistent pain in 7 patients, unexplained fever in 2, perforation in 2, and delayed tracheo-esophageal fistula in 1. Two patients died of treatment-related complications. Repeated dilation was required in 32 of the 75 patients on long-term follow-up. We conclude that adequate palliation of dysphagia can be achieved by dilation in two-thirds of patients with radiation therapy-induced strictures of the esophagus. Dilation of these strictures is relatively simple and safe if performed with care.

Diagnosis of malignant obstructive jaundice by bile cytology: results improved by dilating the bile duct strictures.

The disruption of malignant biliary strictures by dilation could enhance the results of bile cytology. To test this hypothesis, we studied the results of bile cytology in 64 consecutive patients undergoing endoscopic biliary drainage for malignant biliary strictures. Patients included 36 men and 28 women, ages 29 to 79 years. In the control group (n = 15), bile was obtained by aspiration without dilating the biliary stricture. In the dilated group (n = 49), bile was aspirated after dilating the biliary stricture to 10F gauge. The bile was centrifuged, and smears were prepared, stained, and interpreted as malignant, suggestive of malignancy, or not malignant. The biliary obstruction was caused by gallbladder cancer in 33, primary bile duct cancer in 14, pancreatic cancer in 11, and metastasis in 6 patients. Forty patients had obstruction at the bifurcation of the hepatic duct. Malignancy was confirmed by surgery in 14, fine-needle aspiration cytology in 9, presence of metastasis in 19, and a combination of clinical and radiologic studies, endoscopic cholangiopancreatography findings, elevated tumor markers, and follow-up in 22 patients. Bile cytology was positive for malignancy in 4 (26.6%) and 31 (63.3%), suggestive in 1 (6.7%) and 6 (12.2%), and negative in 10 (66.7%) and 12 (24.5%) patients in the control group and the dilated group, respectively (p = 0.028, 95% CI 1.15 and 21.03). Cytology was positive in 73% of gallbladder cancers, 62.5% of bile duct cancers, 40% of pancreatic cancers, and 60% of metastasized cancers after dilation. Two patients had hemobilia, 8 had cholangitis, and 2 had pancreatitis after biliary drainage.(ABSTRACT TRUNCATED AT 250 WORDS)