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Biomedical subjects

R K Sharma

Publications and source records attributed to R K Sharma.

At least 361 records · Page 20Linked to original sources
PubMed1990

Genetic evidence for alpha 2-adrenergic receptor subtypes in rat brain, heart and adrenal gland.

A complementary DNA (cDNA) clone - cA2-47 - corresponding to a new alpha 2-adrenergic receptor subtype has been isolated from a rat brain cDNA library and used as a hybridization probe to scrutinize the alpha 2-receptor poly(A+) RNAs in rat brain, heart and adrenal gland. Hybridization of the 5' half of the coding region of this cDNA at 37 degrees C to rat brain poly(A+) RNA revealed a single band at 5.8 kb as the size of its corresponding mRNA. Under identical hybridization conditions, a human platelet alpha 2-receptor genomic probe failed to hybridize to any rat brain mRNAs. Under lower stringency conditions, hybridization of the full-length cDNA, cA2-47, to selected rat tissue poly(A+) RNA showed the presence of four different sized mRNAs in brain and three in both heart and adrenal gland. Messages of 1.3 kb and 2.1 kb were common in all three tissues (although the band at 2.1 kb was slightly higher in the heart and adrenal gland). A 5.8 kb mRNA was unique to the brain and a slightly higher band at 6.0 kb was consistently present in heart and adrenal gland but was absent in the brain. A fourth message at 3.4 kb was found predominantly in the brain and was either absent or present at very low levels in the other tissues examined.(ABSTRACT TRUNCATED AT 250 WORDS)

Adrenal Gland Neoplasms↗
PubMed1990

Purification and characterization of novel calmodulin-binding protein from cardiac muscle.

A novel protein which represents the most abundant calmodulin-binding protein in bovine heart cytosolic fraction was purified to apparent homogeneity. The purification procedure involved DEAE-Sepharose CL-6B (to remove calmodulin), calmodulin-Sepharose 4B affinity, and Sepharose 6B column chromatographies. This purified calmodulin-binding protein is a highly asymmetric protein with a sedimentation coefficient of approximately 5.0 S and a Stokes radius of about 83.0 A. The molecular weight of the calmodulin-binding protein was determined to be 175,000 from the sedimentation constant and Stokes radius of the protein. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the protein showed a single protein band with an apparent molecular weight of 140,000. The result suggests that the protein is monomeric. Although this molecular weight is similar to that of caldesmon, a known ubiquitous calmodulin-binding protein, the protein did not react with caldesmon-specific antibodies, nor did it display a proteolytic fragmentation pattern similar to that of the former. In addition, caldesmon was found almost exclusively in the particulate fraction in low ionic strength cardiac muscle extract, whereas this protein is purified the soluble fraction.

Animals↗
PubMed1990

Quantitative estimation of Holarrhena antidysenterica bark total alkaloids in crude drugs and in the body fluids of man and rat.

A turbidimetric method was developed for the quantitative estimation of the total alkaloids of kutaj bark (Holarrhena antidysenterica) in crude medicinal preparations and in the body fluids of man and rat. The alkaloids were colloidally precipitated with Dragendorff's reagent as complex salts of potassium iodobismuthate in extremely dilute solutions. The finely subdivided orange-brown precipitate gave a coloured, clear homogeneous suspension in the presence of gum arabic. Optical density of such suspensions changed linearly with the change in alkaloid concentration, when prepared within the standardized experimental conditions that included control of ion concentration and temperature of the reaction mixture. Observations revealed the reversible nature of the alkaloid-reagent reaction. Crude medicinal preparations from three different pharmaceutical sources contained varying concentrations of the alkaloids. Complete recovery of the alkaloids was possible from plasma and urine, while significant amounts of the alkaloids were lost to blood cells and faecal contents in man and rat.

Alkaloids↗
PubMed1990

Contraceptive potentiality of STS-557--a feasibility study in male bonnet monkey (Macaca radiata).

STS-557 (17 alpha-cyanomethyl-17 beta-hydroxy-estra-4, 9(10)-diene-3-one) was administered (i.m.) to two groups (4 in each group) of adult male bonnet monkeys at a daily dose of 10 mg/monkey for 12 weeks (first group) and 5 mg/monkey for 14 weeks (second group). Treatment with the 10 mg dose resulted in a significant decline in the count, motility, acrosin and hyaluronidase activities and the fertilizing ability (zona-free hamster egg penetration assay) of spermatozoa by the 6th week of initiation of treatment. The circulating level of STS-557 was low after one week and increased from the 2nd week of treatment when the serum level of testosterone was significantly reduced. Complete recovery was observed by the 11th week after withdrawal of treatment. The treatment with the 5 mg dose had minor and inconsistent effect on the motility, hyaluronidase and acrosin activity, and the fertilizing ability of spermatozoa in addition to the blood level of testosterone. STS-557 may have the potentiality to be used as a chemical contraceptive in the male but compensation for the reduced level of blood testosterone may be necessary.

Acid Phosphatase↗
PubMed1990

Airway beta-adrenoceptor number in cystic fibrosis and asthma.

1. Maximal binding capacity (Bmax) and the dissociation constant (KD) for the beta-adrenoceptor antagonist 125I-cyanopindol were estimated in membrane preparations of hilar, lobar/main bronchi (level I) and peripheral lung (level II) of grossly normal lungs resected for bronchial carcinoma. The tissue distribution of 125I-cyanopindol-binding sites was assessed by autoradiography of complementary cryostat sections. The data obtained from the resections for carcinoma and bronchiectasis were used as disease controls for comparison with those obtained from patients with cystic fibrosis and asthma. 2. In carcinoma controls, mean Bmax values (+/- SEM) for airway levels I and II were 89 +/- 4 and 133 +/- 6 fmol/mg of protein, respectively (P less than 0.01). The corresponding KD values at each airway level were similar, i.e. 29 +/- 2 and 33 +/- 1 pmol/l, respectively. Autoradiography revealed that there was dense labelling of bronchial and bronchiolar epithelium and most strikingly of the alveolar wall. 3. Compared with carcinoma controls, mean Bmax values in cystic fibrosis were significantly reduced in membrane preparations of both airway levels I and II (P less than 0.01). Autoradiography showed the reduction was most apparent in alveolar wall and bronchial epithelium. 4. There was a tendency to reduction of Bmax in membrane preparations from patients with bronchiectasis at airway level I, but this failed to reach statistical significance. Autoradiography demonstrated that the density of labelling was significantly reduced in bronchial epithelium and bronchial smooth muscle as compared with carcinoma controls (P less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1990

Effect of STS-557 (17 alpha-cyanomethyl 17 beta-hydroxy-estra-4, 9(10)-dien-3-one) on blood hormone levels, the testis, accessory sex organs and fertility of rats.

Treatment with STS-557 (17 alpha-cyanomethyl 17 beta-hydroxy-estra-4,9(10)-dien-3-one; 10 mg kg-1 daily s.c.) for 4 weeks induced atrophy of the seminiferous tubules in adult rats with a reduction in tubule diameter and in the number of round spermatids at stage VII. Elongated spermatids were not detected. Leydig cells were atrophied from the second week of treatment with a concomitant decrease in blood levels of testosterone. The blood levels of FSH and LH were reduced from the third week of treatment. The weight of the reproductive organs was reduced after STS-557 treatment. The treatment induced sterility in 50% of rats after 2 weeks of treatment but after 4 weeks none of the treated males mated. Normal fertility and normal levels of testosterone and FSH were restored after 6 weeks and LH after 4 weeks of withdrawal of treatment. All other parameters studied recovered to pretreatment levels 6 weeks after withdrawal of treatment. STS-557 could act on the pituitary-gonadal axis (reducing gonadotrophin secretion) as well as directly affecting the Leydig cells. The consequent reduction in the blood levels of testosterone in combination with reduced gonadotrophins was presumably responsible for the suppression of spermatogenesis.

Animals↗
PubMed1990

Therapeutic role of carbogen in impaired hearing.

The therapeutic role of carbogen was evaluated in subjects with sensorineural hearing loss by administering carbogen, a gas mixture of 95 per cent O2 and 5 per cent CO2, for seven consecutive days (30 min/day) and monitoring puretone audiometry before and after the administration. Significant improvement was observed both in air and bone conduction threshold levels on seventh day, indicating that there was some recoverable portion in the hearing level of these subjects. The improvement in hearing may be due to action of CO2 as an otic vasodilator coupled with supplementation of the O2 requirement of degenerating hair cells. Carbogen thus appears to be useful in persons with impaired hearing, involving the inner ear.

Adult↗
PubMed1990

Effect of cambendazole and haloxon on the carbohydrate metabolism of Ascaridia galli and Heterakis gallinae (Nematoda).

Cambendazole 10(-4) M significantly reduced the level of glycogen in both the parasites. Lactic acid level was found enhanced. Oxygen consumption was suppressed by 63 and 94% in A. galli and H. gallinae, respectively, by 10(-4) M cambendazole. Haloxon did not cause any significant change in glycogen and lactic acid level in either parasite, but reduced oxygen consumption by 23 and 31% in Ascaridia galli and Heterakis gallinae, respectively.

Animals↗
PubMed1989

Role of hepatic and renal cytochrome P-450 IVA1 in the metabolism of lipid substrates.

The role of clofibrate-inducible cytochrome P-450 IVA1 in the metabolism of endogenous lipids in both rat liver and kidney microsomal fractions has been investigated. 20(omega)-hydroxyarachidonic acid has been identified as a major metabolite after incubation with both tissue fractions and the structure confirmed by mass spectrometry. The arachidonic acid 20-hydroxylase activity is inducible by clofibrate in both liver and kidney, indicating that cytochrome P-450 IVA1 is probably the enzyme responsible for this activity. In addition, the kidney exhibited higher rates of arachidonate 20-hydroxylase activity than the liver (in both control and induced states). Although leukotriene B4 was also hydroxylated in the 20-position in both liver and kidney, clofibrate induction resulted in a decrease (approximately 50%) in hydroxylase activity. In addition, the absolute level of leukotriene B4 20-hydroxylase activity in both tissue homogenates and by purified cytochrome P-450 IVA1 in a reconstituted system, was 2-3 orders of magnitude lower than the corresponding activity for lauric acid and arachidonic acid as substrates, indicating that the leukotriene was not the preferred substrate for this enzyme. Computer modelling of the conformational geometries of the above three potential cytochrome P-450 IVA1 substrates have shown that both lauric and arachidonic acids adopt a compact, 'hairpin' structure that are almost superimposed on each other, thereby rationalizing why they are relatively good substrates for this isoenzyme. By contrast, leukotriene B4 adopts a more bulky geometry than the two fatty acids, thereby providing a coherent structural reason why it is a poorer substrate for the cytochrome P-450 IVA1 isoenzyme.

Animals↗
PubMed1989

Differential induction of peroxisomal and microsomal fatty-acid-oxidising enzymes by peroxisome proliferators in rat liver and kidney. Characterisation of a renal cytochrome P-450 and implications for peroxisome proliferation.

The induction of renal fatty-acid-oxidising enzymes has been investigated following short-term exposure to a group of structurally diverse peroxisome proliferators and compared to the more extensively documented hepatic responses in the rat. There was a marked compound dependence on induction of both cytochrome P-450-IVA1-dependent omega-hydroxylation of lauric acid and enzymes of the peroxisomal fatty acid beta-oxidation pathway (measured as cyanide-insensitive palmitoyl-CoA oxidation and enoyl-CoA hydratase). Cytochrome P-450 IVA1 (or a very closely related isoenzyme in the same gene family) was a major constitutive haemoprotein in rat kidney microsomes and actively supported the omega-hydroxylation of lauric acid. This activity was induced 2-3-fold by peroxisome proliferators such as clofibrate, di-(2-ethylhexyl)phthalate, bezafibrate and nafenopin. By using a cDNA probe to the cytochrome P-450 IVA1 gene in Northern blot analysis, we have shown that increased renal and hepatic omega-hydroxylation of lauric acid, after treatment with peroxisome proliferators is a consequences of a substantial increase in the mRNA coding for this haemoprotein. In addition, programming of an in vitro rabbit reticulocyte translation system with both renal and hepatic RNA resulted in the synthesis of similar (if not identical) cytochrome-P-450-IVA1-related polypeptides. Furthermore, we have provided Western blot evidence that both rat liver and kidney microsomes contain two closely related cytochrome P-450 IVA1 polypeptides, the major one characterised by a monomeric molecular mass of 51.5 kDa (identical to authentic, purified hepatic cytochrome P-450 IVA1) and a minor one of 52 kDa. The kidney-supported fatty acid omega-hydroxylase activity was refractory to inhibition by a polyclonal antibody to liver cytochrome P-450 IVA1, which may be related to the existence of two closely related (but immunochemically distinct) fatty acid hydroxylases in this tissue. Our studies have also demonstrated that certain of the compounds tested (including clofibrate, bezafibrate and nafenopin) induced renal fatty acid beta-oxidation, mirroring the increased omega-hydroxylase activity in the endoplasmic reticulum. Our studies have also indicated that the kidney was more refractory to induction of the endoplasmic reticulum and peroxisomal fatty-acid-oxidising enzymes than the liver. Taken collectively, our data is strongly suggestive of a possible linkage of the renal fatty acid oxidative enzymes in these two organelles, a situation that also occurs in the liver. In addition, our studies have provided a possible conceptual framework that may rationalise the decreased susceptibility of the k

Amino Acid Sequence↗
PubMed1989

Regulation of Ca2+/calmodulin-dependent cyclic nucleotide phosphodiesterase by the autophosphorylated form of Ca2+/calmodulin-dependent protein kinase II.

The 63-kDa subunit, but not the 60-kDa subunit, of brain calmodulin-dependent cyclic nucleotide phosphodiesterase was phosphorylated in vitro by the autophosphorylated form of Ca2+/calmodulin-dependent protein kinase II. When calmodulin was bound to the phosphodiesterase, 1.33 +/- 0.20 mol of phosphate was incorporated per mol of the 63-kDa subunit within 5 min with no significant effect on enzyme activity. Phosphorylation in the presence of low concentrations of calmodulin resulted in a phosphorylation stoichiometry of 2.11 +/- 0.21 and increased about 6-fold the concentration of calmodulin necessary for half-maximal activation of the phosphodiesterase. Peptide mapping analyses of complete tryptic digests of the 63-kDa subunit revealed two major (P1, P4) and two minor (P2, P3) 32P-peptides. Calmodulin-binding to the phosphodiesterase almost completely inhibited phosphorylation of P1 and P2 with reduced phosphorylation rates of P3 and P4, suggesting the affinity change of the enzyme for calmodulin may be caused by phosphorylation of P1 and/or P2. When Ca2+/calmodulin-dependent protein kinase II was added without prior autophosphorylation, there was no phosphorylation of the 63-kDa phosphodiesterase subunit or of the kinase itself in the presence of a low concentration of calmodulin, and with excess calmodulin the phosphodiesterase subunit was phosphorylated only at P3 and P4. Thus the 63-kDa subunit of phosphodiesterase has a regulatory phosphorylation site(s) that is phosphorylated by the autophosphorylated form of Ca2+/calmodulin-dependent protein kinase II and blocked by Ca2+/calmodulin binding to the subunit.

Animals↗
PubMed1989

Molecular comparison of alpha 2-adrenergic receptors from rat adrenocortical carcinoma and human blood platelet.

We have previously described a simple two-step purification technique to isolate alpha 2-adrenergic receptors from the rat adrenocortical carcinoma (Jaiswal, R. K. and Sharma, R. K. (1985) Biochem. Biophys. Res. Commun. 130, 58-64). Utilizing this technique we have now achieved approximately 77,000-fold purification to apparent homogeneity of alpha 2-adrenergic receptors from human platelets. We have compared the biochemical characteristics of these receptors with those from the rat, which were purified approximately 40,000-fold to homogeneity. The [125I] receptor proteins from two sources showed: (a) a single radioactive band with a Mr of 64,000 as evidenced by one- and two-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE); and (b) a single symmetrical peak with a pI of 4.2 by isoelectric focusing polyacrylamide gel electrophoresis. Both proteins showed typical alpha 2-adrenergic binding characteristics with specific binding activities of 13.85 nmol/mg and 14.17 nmol/mg protein. These values are close to the theoretical binding activity of 15.6 nmol/mg protein for 1 mol of the ligand binding 1 mol of the receptor protein. These results attest to the purity of the receptors, to its Mr of 64,000, and to its acidic nature. However, the peptide maps of the radioiodinated alpha 2-adrenergic receptors from rat adrenocortical carcinoma and human blood platelets reveal some distinct differences which may relate to the differences in the pharmacological specificities between rodent and non-rodent alpha 2-adrenergic receptors.

Adrenal Gland Neoplasms↗
PubMed1989

A monoclonal antibody which identifies the autophosphorylation domain of autophosphorylating protein kinase 500.

Autophosphorylating protein kinase 500 is a serine protein kinase expressed progressively with the steps of cellular transformation, approaching levels from 50- to 100-fold in the terminal stages of malignancy. The enzyme possesses a sharply restricted range of substrates: itself and a ribosomal protein with a molecular weight of 31,000 (S6). We report here on the characterization of a monoclonal antibody directed against the autophosphorylation domain of AUT-PK 500. The specificity of the antibody is evidenced by blockage of the enzyme phosphorylation without interfering with the native S6 or synthetic octapeptide (S6-1) serine residue phosphorylations. This represents an important step in identifying a probe that can be used to explore the structure and potential function of AUT-PK 500 in cellular transformation.

Animals↗
PubMed1989

The effect of levamisole and albendazole on some enzymes of Ascaridia galli and Heterakis gallinae.

Ascaridia galli and Heterakis gallinae obtained from the common fowl Gallus gallus were exposed to 10(-2)-10(-5)M levamisole and albendazole; both compounds caused death of the parasites in vitro. The effect of the drugs was investigated on homogenates of the treated worms. Albendazole, at 10(-2)M, inhibited oxaloacetate reduction by 67 and 53% and malate oxidation by 21 and 17% in A. galli and H. gallinae, respectively, whereas 10(-4)M levamisole completely inhibited malate dehydrogenase activity in both directions in the two parasites. Lactate dehydrogenase was not affected significantly by either anthelmintic. Aldolase activity was diminished by 57 and 32% in A. galli and H. gallinae, respectively, with 10(-4)M levamisole. Levamisole at 10(-4)M also inhibited the activity of acid and alkaline phosphomonoesterase and cholinesterase. Albendazole had no significant effect on these enzymes in either parasite. Malate dehydrogenase and cholinesterase activity of the host tissue (intestine and caecum) was also reduced significantly with 10(-2) and 10(-3)M levamisole. These studies indicated a multiple mode of action of levamisole and albendazole.

Albendazole↗
PubMed1989

Long-term experience with 1 per cent topical silver sulphadiazine cream in the management of burn wounds.

Three hundred and forty-two patients with 10-50 per cent body surface area burns were studied prospectively over the 5-year period from 1982 to 1986 for the effectiveness of topical 1 per cent silver sulphadiazine. Various parameters were studied including: (i) healing time of deep partial skin thickness burns, (ii) eschar separation time, (iii) conversion rate of deep dermal burns to full skin thickness burns, (iv) burn wound surface bacterial flora and their changing pattern over the years, (v) incidence of invasive sepsis and (vi) overall mortality. There was a remarkable decrease in the time taken for the healing of deep dermal burns, and the conversion rate of deep dermal burns to full skin thickness was significantly reduced. Eschar separation was delayed considerably. There was a total change in the predominent surface micro-organisms from Staph. aureus, which was predominant in 1982, to pseudomonas species and klebsiella in 1986. Moreover, there was the emergence of a new variety of micro-organism within the last 2 years. The incidence of invasive infection and overall mortality was significantly reduced.

Administration, Topical↗