Biomedical subjects

A Molla

Publications and source records attributed to A Molla.

At least 19 recordsLinked to original sources

PubMed1992

Cardioviral internal ribosomal entry site is functional in a genetically engineered dicistronic poliovirus.

High mutation rates have driven RNA viruses to shorten their genomes to the minimum possible size. Mammalian (+)-strand RNA viruses and retroviruses have responded by reducing the number of cis-acting regulatory elements, a constraint that has led to the emergence of the polyprotein. Poliovirus is a (+)-stranded picornavirus whose polyprotein, encoded by an open reading frame spanning most of the viral RNA, is processed by virus-encoded proteinases. Despite their genetic austerity, picornaviruses have retained long 5' untranslated regions, which harbour cis-acting elements that promote initiation of translation independently of the uncapped 5' end of the viral messenger RNA. These elements are termed 'internal ribosomal entry sites' and are formed from highly structured RNA segments of at least 400 nucleotides. How these elements function is not known, but special RNA-binding proteins may be involved. The ribosome or its 40S subunit probably binds at or near a YnXmAUG motif (where Y is a pyrimidine and X is a purine) at the 3' border of the internal ribosomal entry site, which either provides the initiating codon or enables the ribosome to translocate to one downstream (E.W. et al., submitted). Initiation from most eukaryotic messenger RNAs usually occurs by ribosomal recognition of the 5' and subsequent scanning to the AUG codon. Here we describe a genetic strategy for the dissection of polyproteins which proves that an internal ribosomal entry site element can initiate translation independently of the 5' end.

Binding Sites↗

PubMed1992

Lack of transcription and expression of the alpha IIb integrin in human early haematopoietic stem cells.

The glycoprotein IIb, the alpha subunit of the platelet integrin GPIIb-IIIa, is a marker of megakaryocyte, but the stage of its expression during haematopoiesis remains controversial. We have examined the expression of GPIIb protein and alpha IIb mRNA in early human normal stem cells. We have purified stem cell expressing the CD34 surface marker (CD34+ fraction) and selected among this population quiescent cells (CD34+ MF(R) fraction). We have failed to detect GPIIb protein and alpha IIb mRNA in the pluripotential (CD34+ MF(R)) cells, even with polymerase chain amplification. Therefore alpha IIb transcription and GPIIb protein expression seemed to follow the commitment of the pluripotential cell in the megakaryocyte lineage.

Antigens, CD↗

PubMed1992

Mutational analysis of the proposed FG loop of poliovirus proteinase 3C identifies amino acids that are necessary for 3CD cleavage and might be determinants of a function distinct from proteolytic activity.

Mutations were introduced into a cDNA clone of poliovirus resulting in single-amino-acid substitutions within the region of the proposed FG loop of proteinase 3C. RNAs were made by in vitro transcription with T7 RNA polymerase and used to transfect HeLa cells. Virus viability was assessed as indicated by cell lysis. In parallel, RNAs were translated in vitro by using a HeLa cell lysate, and the patterns of the processed poly-proteins were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Replacement of Lys-78, Arg-79, and Glu-81 had apparently no effect on virus viability and on proteolytic processing. In contrast, virus viability was abolished by mutation of Phe-83, Arg-84, Asp-85, Ile-86, and Arg-87. With respect to substitution of Phe-83, Asp-85, and Arg-87, these effects correlated with impaired processing of the 3CD cleavage site, separating 3C and 3D, and, to a lesser extent, of the P1 precursor. Replacement of Arg-84 and Ile-86, on the other hand, did not alter the processing pattern. Thus, the lethal effects in these mutant genomes may not have been caused by impaired processing. A special case was the mutant of Lys-82-Gln. Virus recovered from cells transfected with RNA carrying this mutation always contained an A-to-G transition which resulted in the replacement of glutamine for arginine. Our data suggest that residues in the proposed FG loop of proteinase 3C influence 3CD cleavage and that they are determinants of a function unrelated to proteolytic processing.

3C Viral Proteases↗

PubMed1992

Prevalence of iron deficiency anaemia in children of the urban slums of Karachi.

A preliminary baseline survey was conducted to estimate the prevalence of anaemia in a group of 391 children aged 6-60 months, randomly selected from three urban slums of Karachi. Haemoglobin and the red cell indices including haematocrit, MCV, MCH, MCHC, RBC and red cell distribution width (RDW) were estimated for each of the selected children. Ferritin estimation was done on 354 (91%) children to assess the iron storage status. According to WHO criteria, the accepted cut-off point for anaemia screening in children is set at 11 gm/dl, 70 fl and 20 pg for haemoglobin, MCV and MCH respectively. Following these criteria, 118 (30%) children were classified as normals (Hg = greater than 11 gm/dl) and 273 (70%) as anaemic (Hg = less than 11 gm/dl). Of the 354 ferritin estimations, 225 (64%) children had ferritin levels lower than normal (less than 11 ng/ml) and 128 (36%) had ferritin levels within normal limit (11-120 ng/ml). From this group, a total of 61% (214/354) children were classified as microcytic hypochromic (MIH) and 11% (39/354) of which had normal ferritin levels suggesting the presence of thalassemia minor trait. The overall results obtained indicate that iron deficiency anaemia is highly prevalent among these children.

Anemia, Hypochromic↗

PubMed1991

Cell-free, de novo synthesis of poliovirus.

Cell-free translation of poliovirus RNA in an extract of uninfected human (HeLa) cells yielded viral proteins through proteolysis of the polyprotein. In the extract, newly synthesized proteins catalyzed poliovirus-specific RNA synthesis, and formed infectious poliovirus de novo. Newly formed virions were neutralized by type-specific antiserum, and infection of human cells with them was prevented by poliovirus receptor-specific antibodies. Poliovirus synthesis was increased nearly 70-fold when nucleoside triphosphates were added, but it was abolished in the presence of inhibitors of translation or viral genome replication. The ability to conduct cell-free synthesis of poliovirus will aid in the study of picornavirus proliferation and in the search for the control of picornaviral disease.

Base Sequence↗

PubMed1991

Activation of ras oncogenes and expression of tumor-specific transplantation antigens in methylcholanthrene-induced murine fibrosarcomas.

The DNA of 22 fibrosarcomas, newly induced in BALB/c mice by subcutaneous doses of 3-methylcholanthrene (3-MCA), was tested in NIH 3T3 transformation assay. Activation of K-ras and N-ras was found in 7 and 3 cases respectively. No H-ras activation was detected. Polymerase chain reaction and oligonucleotide hybridization performed on the DNA of the 22 sarcomas revealed 5 cases of K-ras mutation at codon 12, 3 at codon 13 and 1 at both codons. One case of K13 mutation was not detectable by transfection. Three cases of mutation at codon 61 of N-ras were also found, one of which was simultaneous with a K12 mutation. Tumor-specific transplantation antigens (TSTA) were assessed in the 22 original tumors. Altogether 16 sarcomas were immunogenic, with the highest frequency of TSTA+ tumors (10/11 and 5/6) in the groups given 1.0 and 0.1 mg of 3-MCA respectively, the lowest (1/5) in that with 0.01 mg of carcinogen; ras mutations occurred in the DNAs of 11 out of the 16 TSTA+ sarcomas, but none of the DNAs of the 6 TSTA- tumors showed ras mutation. The results suggest that 3-MCA-induced transformation of subcutaneous fibroblasts can involve mutations in codons 12, 13 or 61 of K- and N- but not H-ras gene and that such mutation is accompanied by the expression of TSTA.

Animals↗

PubMed1991

Factors associated with elevated serum cholesterol levels in well-to-do Pakistani schoolchildren.

Cholesterol (CH) and triglyceride (TG) levels were determined in blood drawn after an overnight fast from 388 school children aged 5-19 years from private schools in Karachi. The mean CH levels ranged from 4.4 to 4.6 mmol l-1 (170.1 to 177.9 mg dl-1) for boys and 4.4 to 4.8 mmol l-1 (170.1 to 185.6 mg dl-1) for girls. The range of TG levels was 1.0 to 1.2 mmol l-1 (88.6 to 106.3 mg dl-1) and 0.9 to 1.1 mmol l-1 (79.7 to 97.4 mg dl-1) for boys and girls respectively. Sixty-two per cent of the girls and 54% of the boys had cholesterol values greater than or equal to 4.4 mmol l-1 (170 mg dl-1), a level at which dietary intervention is recommended for children. Thirty-two per cent of all the children had triglyceride levels above the 90th percentile of the levels for similar age groups in North America. The mean cholesterol intake was 469 mg/day for girls and 518 mg/day for boys. Overweight and inactivity were associated with raised serum cholesterol levels. Forty per cent of the girls and 25% of the boys reported a strong family history of hypercholesterolaemia and/or heart disease. The results show that the prevalence of hypercholesterolaemia is high in well-to-do Pakistani school children and factors which can be modified to lower serum cholesterol levels are identified.

Adolescent↗

PubMed1990

Immunogenicity and antigenicity of conserved peptides from the envelope of HIV-1 expressed at the surface of recombinant bacteria.

We expressed peptides from the HIV-1 envelope protein at the surface of Escherichia coli by genetic insertions into an exposed loop of the outer membrane protein LamB. Recombinant bacteria expressing eight peptides from gp110 (pep1-pep8), conserved between HIV-1 and HIV-2, were used as live immunogens in rabbits by the intravenous route. The eight constructions elicited anti-LamB antibodies, showing that the hybrid proteins were immunogenic. One of them, LamB-pep8, gave rise to antibodies able to react with gp160 and to neutralize HIV-1 in vitro. We also show that this type of recombinant E. coli can provide a convenient reagent to monitor and characterize specific antibodies. Recombinant clones were used to test sera of seropositive individuals, as well as to narrow down the monoclonal antibody 110-1 recognition site to a cluster of eight residues at the carboxy-terminal end of gp110.

Amino Acid Sequence↗

PubMed1990

Inactivation of chemotactic activity of C5a by the serratial 56-kilodalton protease.

The effects of the 56-kilodalton protease (56K protease) from Serratia marcescens on complement-derived chemotactic activity were examined. Fresh human serum was incubated with zymosan to produce C5a. This activated serum was then incubated with various concentrations of 56K protease, and the chemotactic activity of mouse peritoneal exudate polymorphonuclear leukocytes (PMN) and macrophages was evaluated. A significant dose-dependent decrease of chemotactic activity was observed after protease treatment. Furthermore, treatment of human recombinant C5a with 56K protease at a dose of 1.0 microgram/ml resulted in a complete loss of chemotactic activity. When the living bacteria of the virulent strain, which produced about 10 times more protease than did the less virulent strain, were injected intraperitoneally into mice, the magnitude of infiltration of polymorphonuclear leukocytes into the peritoneal cavity was much lower than that caused by the less virulent strain. Because complement-dependent chemotactic activity is an initial response to bacterial infection, these results suggest indirect pathogenic functions of serratial proteases that suppress chemotactic activity.

Blood Bactericidal Activity↗

PubMed1990

Serum cholesterol in neonates and their mothers. A pilot study.

Reports from the Aga Khan University indicate that 58% of 400 school children studied had undesirably high serum cholesterol levels. The present study was undertaken to determine whether the high cholesterol levels are present at birth and to determine the relationship between cord blood, maternal blood cholesterol and maternal diet. Cord blood from 58 neonates and fasting venous blood from 45 mothers were analyzed from total serum cholesterol. Mothers were interviewed regarding their usual diet during pregnancy. Mean cord blood cholesterol was 56.90 mg/dl (range 26 to 123 mg/dl). Mean maternal blood cholesterol was 232.4 mg/dl (range 141-382 mg/dl). Mean maternal intake of cholesterol was 457 mg (recommended level less than or equal to 300 mg/day). There was no significant co-relation between cord blood cholesterol and maternal blood cholesterol or maternal intake of cholesterol. Eighteen percent of the mothers reported a strong family history of hypercholesterolemia and/or heart disease, but this genetic tendency was not observed in the blood cholesterol level at birth indicating that environmental factors namely diet may have a prime role in determining serum cholesterol levels in childhood.

Cholesterol↗

PubMed1990

Serum alkaline phosphatase in apparently healthy Karachi population.

Serum alkaline phosphatase (AP) was estimated in a total of 786 (418 males and 386 females) apparently healthy people aged between 1-75 years selected randomly from a sample of the Karachi population. Reference ranges for AP level were obtained for the males and females stratified into ten successive age groups. The study population was also divided into two main age groups, a paediatric aged between 1-14 years and an adult group aged between 15 to over 50 years to see if the mean AP levels differ significantly between the two groups. Mean AP level for the male paediatric age group was 225 IU/L, significantly higher (P less than 0.005) than those of the male adult mean level of 83 IU/L. Similarly the mean AP levels for the paediatric female age group was 205 IU/L significantly higher (P less than 0.005), compared to the mean AP level of 67 IU/L obtained for the female adult age group.

Adolescent↗

PubMed1990

Role of glucose polymer (cereal) in oral rehydration therapy.

The standard packaged glucose-based oral rehydration solution (ORS) provides optimal rehydration of acute diarrhea from any cause, but it does not reduce the volume, frequency, or duration of diarrhea. A new ORS formulation has been developed in which glucose is replaced by 50 to 60 gm of cereal flours, such as rice, wheat, maize, sorghum, or millet, or equivalent amounts of noncereal staples, such as boiled potato or plantain. In a clinical trial in children suffering from acute diarrhea, the staple-based or polymer-based ORS achieved a 40% to 60% reduction in the stool volume compared with the standard ORS. A three-cell longitudinal study in rural Bangladesh involving 2,000 children aged 1 to 4 years demonstrated the superior efficacy of rice ORS compared with glucose ORS or no ORS. The cumulative recovery rate on day 3 was 66%, 24%, and 11% in the rice ORS, glucose ORS, and comparison groups, respectively. The study suggests that staple-based or food-based ORS is the optimal treatment of diarrhea.

Diarrhea↗

PubMed1990

Blood lipids in a healthy Karachi population.

Serum levels for cholesterol and triglycerides were estimated in an apparently normal healthy population of Karachi, aged between 4 and 59 years. In total, there were 632 subjects, 322 males and 310 females. Hypercholesterolemia was defined as a cholesterol level greater than 6.2 mmol l-1 (240 mg dl-1) in subjects above 20 years of age. In the age groups 20-39 and 40-59 years hypercholesterolemia was present in 26-41% of the males and 10-38% of the females. When triglyceride levels of more than 2.8 mmol l-1 (250 mg dl-1) were taken as abnormal for healthy males and females, 0-2% of the females and 10-25% of the males above 20 years of age were hypertriglyceridemic. The mean cholesterol levels in the age groups 4-9 and 10-19 years varied from 4.4 to 4.9 mmol l-1 (169.8 to 189.1 mg dl-1).

Adolescent↗

PubMed1989

Pathogenic potentials of bacterial proteases.

Six separate molecular mechanisms for pathogenesis attributed to bacterial proteases are described. (I). Enhancements of vascular permeability and edema formation which result from the activation of kinin generating cascade such as Hageman factor by the proteases. (II). Degradation of defense oriented proteins including IgG and IgA as well as destruction of structural matrices such as fibronectin, proteoglycan and collagen. (III). Inactivation of complement system and generated chemotactic factor from C3 and C5. (IV). Degradation of regulatory plasma protease inhibitors (serpins) including alpha 1-protease inhibitor, alpha 2-macroglobulin (alpha 2M), C1-esterase inhibitor, alpha 2-antiplasmin and antithrombin-III. (V). The protease forms a transitory stable enzyme/inhibitor(alpha 2M) complex. It binds to and internalizes into the cells which possess alpha 2M-receptor such as fibroblasts via the alpha 2M-receptor, and the protease activity is regenerated in cells, and subsequently intracellular integrity is destroyed resulting in cell killing. (VI). The serratial 56 kDa (56K) protease is found to potential viral yield 100 fold more when influenza virus infected mice were subjected to administrations of this protease intranasally. This results in rapid and much elevated lethality.

Animals↗

PubMed1989

Antibodies against synthetic peptides and the topology of LamB, an outer membrane protein from Escherichia coli K12.

LamB, an outer membrane protein from Escherichia coli K12, is involved in the transport of maltose and maltodextrins across the outer membrane and constitutes a receptor for a number of bacteriophages. A recent folding model proposes that LamB spans the outer membrane through a number of transmembranous segments separated by regions exposed either to the cell exterior or to the periplasm. This model is essentially based on predictions of structure and genetic arguments relying on the hypothesis that the mutations studied did not alter the folding of the protein. In order to obtain direct evidence with the unaltered protein, we elicited polyclonal antibodies against synthetic peptides corresponding to several LamB sequences. We chose four regions. Three of them [aa 147-161 (peptide 2), aa 371-385 (peptide 3), and aa 399-413 (peptide 4)] are predicted to face the outside of the cell, and the fourth (aa 19-33 (peptide 1)] is predicted to be periplasmic. By immunoblotting against extracts of various mutants, these antibodies were shown to be specific for LamB and targeted to the selected regions. In some cases, the recognition sites for antibodies were narrowed down to parts of a region. In vivo, on intact cells, anti-peptides 2, 3, and 4 reacted with LamB in an ELISA; this confirmed that regions of peptide 2 and 3 are located, at least in part, at the cell exterior and provided the first proof for a similar, situation of the region of peptide 4. Under the same conditions, anti-peptide 1 did not react with LamB.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

PubMed1989

Food-based oral rehydration salt solution for acute childhood diarrhoea.

The efficacy in acute childhood diarrhoea of oral rehydration therapy (ORT) based on staple foods (maize, millet, wheat, sorghum, rice, or potato) was compared with that of standard ORT based on glucose. 266 children aged 1-5 years, with a history of acute diarrhoea for 48 h or less, moderate to severe dehydration, and no complications, were assigned to treatment with one of the food-based oral rehydration salt solutions (ORS) or standard ORS. The mean stool output over the first 24 h of treatment in the group receiving standard ORS was significantly higher than that of any other treatment group, and the groups receiving food-based ORT showed substantial reductions in stool output compared with the standard ORT group. Abnormalities in electrolyte concentrations were corrected in all treatment groups with similar efficiency. The digestibility of the food-based ORS was assessed by the stool pH, glucose content before and after acid hydrolysis, and osmolality; there were no significant differences between the standard ORS and food-based ORS groups. Food-based ORT should be more acceptable to users in developing countries since the mixtures are similar to traditional weaning foods and since, unlike standard ORT, it reduces stool output substantially.

Acute Disease↗

PubMed1989

Activation of hageman factor and prekallikrein and generation of kinin by various microbial proteinases.

Activation of the Hageman factor-kallikrein-kinin system by serratial 56-kDa proteinase was previously demonstrated (Matsumoto, K., Yamamoto, T., Kamata, T., and Maeda, H. (1984) J. Biochem. (Tokyo) 96, 739-749; Kamata, R., Yamamoto, T., Matsumoto, K., and Maeda, H. (1985) Infect. Immun. 48, 747-753). To investigate whether the activation of the system is specific for 56-kDa proteinase or is found similarly with other microbial proteinases, 11 proteinases of microbial origins were studied; the 56-kDa proteinase was the control. For in vitro studies, activation of guinea pig Hageman factor and prekallikrein was examined in purified systems as well as in plasma as a zymogen source. Specific antibodies and inhibitors confirmed the activation steps of the cascade. In the in vivo study the enhancement of vascular permeability in guinea pig skin and its sensitivity to inhibitors of activated Hageman factor, plasma kallikrein, or a kininase were examined. The results from the in vivo experiments were consistent with those in vitro. Taking all the data together, we classified the 11 microbial proteinases into three groups as follows: 1) Serratia marcescens 56-, 60-, and 73-kDa proteinases, Pseudomonas aeruginosa alkaline proteinase and elastase, and Aspergillus melleus proteinase (this group activated Hageman factor but not prekallikrein); 2) Vibrio vulnificus proteinase, subtilisin from Bacillus subtilis, and thermolysin from Bacillus stearothermophilus (this group activated both Hageman factor and prekallikrein); 3) Streptomyces caespitosus proteinase and V8 proteinase from Staphylococcus aureus (this group activated neither Hageman factor nor prekallikrein, but generated kinin from high molecular weight kininogen directly).

Animals↗

PubMed1989

Cytotoxicity of bacterial proteases in various tumor cells mediated through alpha 2-macroglobulin receptor.

The binding and cytotoxicity of a complex of fluorescein isothiocyanate-labeled 56K protease and alpha 2-macroglobulin (alpha 2M) were determined by using various human and rodent tumor cell lines. The binding was higher at 37 degrees C than at 4 degrees C; a rapid and progressive uptake that was time dependent was noted at 37 degrees C, whereas no uptake was observed at 4 degrees C, which indicated temperature-dependent internalization. The binding was highest in the fibroblastic and adenocarcinoma cells, and lowest in squamous and epidermoid cells. The Scatchard plots for the binding isotherms were linear, with an apparent Kassoc 1.17 to 2.99 x 10(-8) M for those cells with high alpha 2M receptor. The number of binding sites (alpha 2M receptor) per cell was 1.3 to 4.75 x 10(6). Values for squamous/epidermoid cells were much lower or undetectable. Fluorescent antibody staining indicated that MCF-7 and other cells with alpha 2M receptor internalized the protease-alpha 2M complex, whereas B-16 melanoma, which has little alpha 2M receptor on the cell surface, did not. Furthermore, when the cytotoxicity of this complex was compared with that of different cell lines, the cells with high rates of uptake of the complex required only a low concentration of the protease and vice versa. These results suggest a possible mechanism of cytotoxic action of protease: alpha 2M receptor-mediated endocytosis of the complex followed by destruction of cellular integrity after regeneration of proteolytic activity. Thus, cells with more alpha 2M receptor require only a low dose for cytotoxic action when compared with cells with little alpha 2M receptor.

Adenocarcinoma↗