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A Mercado

Publications and source records attributed to A Mercado.

At least 19 recordsLinked to original sources
PubMed2001

Functional and molecular characterization of the K-Cl cotransporter of Xenopus laevis oocytes.

The K-Cl cotransporters (KCCs) have a broad range of physiological roles, in a number of cells and species. We report here that Xenopus laevis oocytes express a K-Cl cotransporter with significant functional and molecular similarity to mammalian KCCs. Under isotonic conditions, defolliculated oocytes exhibit a Cl(-)-dependent (86)Rb(+) uptake mechanism after activation by the cysteine-reactive compounds N-ethylmaleimide (NEM) and mercuric chloride (HgCl(2)). The activation of this K-Cl cotransporter by cell swelling is prevented by inhibition of protein phosphatase-1 with calyculin A; NEM activation of the transporter was not blocked by phosphatase inhibition. Kinetic characterization reveals apparent values for the Michaelis-Menten constant of 27.7 +/- 3.0 and 15.4 +/- 4.7 mM for Rb(+) and Cl(-), respectively, with an anion selectivity for K(+) transport of Cl(-) = PO(4)(3-) = Br(-) > I(-) > SCN(-) > gluconate. The oocyte K-Cl cotransporter was sensitive to several inhibitors, including loop diuretics, with apparent half-maximal inhibition values of 200 and 500 microM for furosemide and bumetanide, respectively. A partial cDNA encoding the Xenopus K-Cl cotransporter was cloned from oocyte RNA; the corresponding transcript is widely expressed in Xenopus tissues. The predicted COOH-terminal protein fragment exhibited particular homology to the KCC1/KCC3 subgroup of the mammalian KCCs, and the functional characteristics are the most similar to those of KCC1 (Mercado A, Song L, Vazquez N, Mount DB, and Gamba G. J Biol Chem 275: 30326--30334, 2000).

Amino Acid Sequence↗
PubMed2001

Thiazide-sensitive cotransporter mRNA expression is not altered in three models of hypertension.

BACKGROUND/AIMS: Several lines of evidence support that the kidney is involved in the increase of arterial blood pressure, and some genetic studies suggest that the thiazide-sensitive Na+:Cl- cotransporter could be implicated in the development of hypertension. In the present study, we analyzed the Na+:Cl- cotransporter mRNA levels in the kidney during the development of hypertension in three experimental models. METHODS: The first model included 18 spontaneously hypertensive rats studied at 4, 10, and 16 weeks of age. The second model included 28 Wistar rats with two-kidney, one-clip Goldblatt hypertension studied at 7, 14, 21, and 28 days. The third model included 6 Wistar rats treated with N(G)-nitro-L-arginine methyl ester during 10 days. Respective controls were studied for all models. At the end of each experimental period, the systolic blood pressure was measured in the tail by plethysmography. Individual renal cortex total RNA was extracted, and the mRNA levels of the thiazide-sensitive Na+:Cl- cotransporter were assessed following a semiquantitative RT-PCR strategy. RESULTS: All experimental models developed systemic hypertension. However, the level of mRNA expression of the Na+:Cl- cotransporter did not change in any of the models studied as compared with their respective controls. CONCLUSION: Our results suggest that a change in mRNA levels of the thiazide-sensitive Na+:Cl- cotransporter is not associated with the development of hypertension in spontaneously hypertensive rats, in rats with renovascular hypertension, nor in rats with hypertension induced by nitric oxide synthesis inhibition.

Animals↗
PubMed2000

Functional comparison of the K+-Cl- cotransporters KCC1 and KCC4.

The K(+)-Cl(-) cotransporters (KCCs) are members of the cation-chloride cotransporter gene family and fall into two phylogenetic subgroups: KCC2 paired with KCC4 and KCC1 paired with KCC3. We report a functional comparison in Xenopus oocytes of KCC1 and KCC4, widely expressed representatives of these two subgroups. KCC1 and KCC4 exhibit differential sensitivity to transport inhibitors, such that KCC4 is much less sensitive to bumetanide and furosemide. The efficacy of these anion inhibitors is critically dependent on the concentration of extracellular K(+), with much higher inhibition in 50 mm K(+) versus 2 mm K(+). KCC4 is also uniquely sensitive to 10 mm barium and to 2 mm trichlormethiazide. Kinetic characterization reveals divergent affinities for K(+) (K(m) values of approximately 25.5 and 17.5 mm for KCC1 and KCC4, respectively), probably due to variation within the second transmembrane segment. Although the two isoforms have equivalent affinities for Cl(-), they differ in the anion selectivity of K(+) transport (Cl(-) > SCN(-) = Br(-) > PO(4)(-3) > I(-) for KCC1 and Cl(-) > Br(-) > PO(4)(-3) = I(-) > SCN(-) for KCC4). Both KCCs express minimal K(+)-Cl(-) cotransport under isotonic conditions, with significant activation by cell swelling under hypotonic conditions. The cysteine-alkylating agent N-ethylmaleimide activates K(+)-Cl(-) cotransport in isotonic conditions but abrogates hypotonic activation, an unexpected dissociation of N-ethylmaleimide sensitivity and volume sensitivity. Although KCC4 is consistently more volume-sensitive, the hypotonic activation of both isoforms is critically dependent on protein phosphatase 1. Overall, the functional comparison of these cloned K(+)-Cl(-) cotransporters reveals important functional, pharmacological, and kinetic differences with both physiological and mechanistic implications.

Animals↗
PubMed1999

Cloning and characterization of KCC3 and KCC4, new members of the cation-chloride cotransporter gene family.

The K+-Cl- cotransporters (KCCs) belong to the gene family of electroneutral cation-chloride cotransporters, which also includes two bumetanide-sensitive Na+-K+-2Cl- cotransporters and a thiazide-sensitive Na+-Cl- cotransporter. We have cloned cDNAs encoding mouse KCC3, human KCC3, and human KCC4, three new members of this gene family. The KCC3 and KCC4 cDNAs predict proteins of 1083 and 1150 amino acids, respectively. The KCC3 and KCC4 proteins are 65-71% identical to the previously characterized transporters KCC1 and KCC2, with which they share a predicted membrane topology. The four KCC proteins differ at amino acid residues within key transmembrane domains and in the distribution of putative phosphorylation sites within the amino- and carboxyl-terminal cytoplasmic domains. The expression of mouse KCC3 in Xenopus laevis oocytes reveals the expected functional characteristics of a K+Cl- cotransporter: Cl--dependent uptake of 86Rb+ which is strongly activated by cell swelling and weakly sensitive to furosemide. A direct functional comparison of mouse KCC3 to rabbit KCC1 indicates that KCC3 has a much greater volume sensitivity. The human KCC3 and KCC4 genes are located on chromosomes 5p15 and 15q14, respectively. Although widely expressed, KCC3 transcripts are the most abundant in heart and kidney, and KCC4 is expressed in muscle, brain, lung, heart, and kidney. The unexpected molecular heterogeneity of K+-Cl- cotransport has implications for the physiology and pathophysiology of a number of tissues.

Amino Acid Sequence↗
PubMed1998

Editing status of mat-r transcripts in mitochondria from two plant species: C-to-U changes occur in putative functional RT and maturase domains.

The intronic mat-r ORF encodes a protein with significant homology to retroviral reverse transcriptases. Here, we describe the nucleotide sequence of potato mat-r and study the editing status of mat-r transcripts in two systems, potato and wheat, where the mat-r ORF is part of the trans-introns but in two different configurations relative to nad1 exons d and e. In potato and wheat, 13 and 15 C-to-U transitions respectively were observed. Most transcripts were partially edited, but potato transcripts were edited more efficiently than wheat transcripts. As in functional mitochondrial genes, RNA editing increased the similarity between plant mat-r proteins and their homologous non-plant counterparts. Interestingly, editing of mat-r was clustered in the reverse-transcriptase (RT) and the maturase (X) domains, two well defined regions having known functions in other systems. These results, together with the integrity and sequence conservation of mat-r, strongly suggest that the encoded protein plays a functional role in plant mitochondria.

Amino Acid Sequence↗
PubMed1998

Electroneutral Na-coupled cotransporter expression in the kidney during variations of NaCl and water metabolism.

The purpose of the present study was to analyze the long-term regulation of renal bumetanide-sensitive Na+-K+-2Cl- cotransporter and thiazide-sensitive Na+-Cl- cotransporter gene expression during changes in NaCl and water metabolism. Male Wistar rats exposed to high or low NaCl intake, saline loading, dehydration, water loading, and furosemide administration during 7 days were studied. Control groups had access to regular food and tap water. Rats were kept in metabolic cages for 4 days before and during the experiment to determine daily urinary electrolyte excretion and osmolarity. At the end of the experiment, creatinine clearance and serum electrolyte levels were also measured. Kidneys were excised and macroscopically subdivided into cortex and outer and inner medulla. Total RNA was extracted from each individual cortex or outer medulla by use of the guanidine/cesium chloride method. The Na+-K+-2Cl- cotransporter expression in outer medulla total RNA was assessed by nonradioactive Northern blot analysis and the Na+-Cl- cotransporter expression in renal cortex total RNA was assessed by semiquantitative polymerase chain reaction. Experimental maneuvers were adequately tolerated, and all groups developed the appropriate renal response to each challenge. However, the level of expression of both cotransporters did not change in any model, except for a 2.8-fold increase in the Na+-Cl- cotransporter expression during dehydration. We conclude that nephron adaptation to 7-day modifications in NaCl and water metabolism does not include changes in the amount of electroneutral sodium-coupled cotransporter gene expression at the mRNA level.

Animals↗
PubMed1998

[Lipoxygenase and trypsin inhibitor inactivation in soymilk processing by direct milling and ultra high temperature (DM-UHT)].

Soymilk production by a new process based on Direct Milling of soy grits and Ultra High Temperature (DM-UHT) has been studied at laboratory scale in order to evaluate solids and protein extraction, lipoxygenases (LO) and Trypsin Inhibitors (TI) inactivation during milling and heat-treatment steps. For TI measurements in soy extracts and soymilk a modification of the classical Kakade method (17) was used. Highest extraction yields were accomplished at 70 degrees C and 2 minutes milling of soy grits. LO was appreciably inactivated when using, as dispersing medium for milling, 0.01M sodium carbonate (Residual Activity 14%) instead of water (Residual Activity 46%), so in this way lower levels of undesired substances can be generated. LO destruction in the resulting suspension was finished by a short heating (30 seconds) from 70 degrees C to boiling temperature (96 degrees C). On the other hand, TI were not fully inactivated in milling nor even in the steaming step at 96 degrees C for many minutes. The TI were inactivated to the accepted levels for soymilk in the final UHT step at 135 degrees C and 2 minutes, being possible at the same time to carry out the simultaneous microorganisms destruction.

Food Handling↗
PubMed1995

Nephrotic syndrome in autosomal dominant polycystic kidney disease.

Urinary protein excretion is generally less than 1 g/24 h in autosomal dominant polycystic kidney disease (ADPKD), and the association of the nephrotic syndrome with this condition is considered rare. A patient with ADPKD associated with nephrotic-range proteinuria is described. She exhibited a relatively rapid impairment of her renal function. An open renal biopsy revealed focal segmental glomerulosclerosis (FGS) with features consistent with secondary FGS. Twenty-one patients with ADPKD and nephrotic syndrome were retrieved from the literature. Fourteen of them (including this case) had a histopathologic evaluation, and FGS was the dominant diagnoses (five patients). Next in frequency were minimal-change disease and membranous nephropathy, with two patients each. Five other patients had a variety of diagnoses. Thus, it is difficult to ascertain if these associations are coincidental or represent a specific pathogenetic relationship. The evaluation of the data also suggests that the presence of proteinuria and nephrotic syndrome accelerates the course of ADPKD toward ESRD.

Aged↗
PubMed1994

Molecular genetic prenatal diagnosis of congenital adrenal hyperplasia due to 21-hydroxylase deficiency by allele-specific hybridization.

The feasibility and accuracy of gene-specific molecular genetic diagnosis for congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency were studied in a group of 24 pregnancies at 25% risk of carrying an affected fetus. Chorionic villus sampling was performed in the majority of cases. Southern blot analysis was carried out to identify deletions or other gross rearrangements. In parallel, the polymerase chain reaction (PCR) was performed, followed by allele-specific oligonucleotide hybridization (ASO) for a panel of nine known mutations. Mutations were identified in 95% of the chromosomes examined. The molecular diagnosis was accurate in 23 of 24 infants. The most common mutation identified was an A-to-G transition in the second intron (52% of affected chromosomes), the result of an apparent gene conversion. One fetus carried homozygous deletion of CYP21, which accounted for 13% of all affected chromosomes. Other mutations identified included an 8-bp deletion in the third exon (22%); Ile172 to Asn, a nonconservative substitution, in the fourth exon (9%); and Gln318 to term, a nonsense mutation, in the eight exon (4%). No mutation was detected in CYP21 in 5% of obligate-affected chromosomes examined by these methods.

Adrenal Hyperplasia, Congenital↗
PubMed1994

A prospective evaluation of antituberculosis therapy in patients with human immunodeficiency virus infection.

The purpose of this study was to determine the efficacy and toxicity of a standard antituberculosis regimen in patients with human immunodeficiency virus (HIV) infection. We prospectively evaluated 89 patients with tuberculosis and HIV infection at an urban medical center. Eighty-two patients received isoniazid, rifampin, and pyrazinamide, with or without ethambutol, for 2 mo, followed by isoniazid and rifampin for 7 mo. Seven patients received other regimens because of drug resistance or intolerance. Therapy was self-administered in 57 patients and directly observed in 32 cases. All patients showed rapid clinical improvement during the first month of therapy, and sputum cultures reverted to negative after 3 mo in 52 of 54 patients from whom specimens were obtained. Adverse reactions to isoniazid or rifampin prompted alterations in antituberculosis regimens in five patients (6%). Forty patients (45%) died during follow-up, and tuberculosis was a potential contributory cause of death in three cases. Treatment failure occurred in five patients (6%), four of whom were noncompliant with therapy. The fifty patient had an isoniazid-resistant organism. No relapses occurred in 916 patient-months of follow-up posttreatment. We thus conclude that the 9-mo regimen used for treatment of drug-susceptible tuberculosis in HIV-infected patients is effective and well tolerated.

AIDS-Related Opportunistic Infections↗
PubMed1992

[Computed tomography guided fine needle aspiration biopsies].

Computed tomography (CT) allows the performance of fine needle aspiration biopsies in situations in which ultrasound or conventional X rays do not correctly visualize the lesion or the needle tract. One hundred four patients with a clinical suspicion of neoplasia were subjected to CT guided percutaneous aspiration biopsies in a lapse of 6 years. The most frequently biopsied organs were the lung (39.5%), liver and gallbladder (20.2%) and pancreas (15.4%). Forty nine percent of procedures were performed in the thorax and 51% in the abdomen. Of 51 patients with suspicion of thoracic cancer, there was only one false negative and no false positive results, with an overall accuracy of 98%. For abdominal procedures, there were no false positive results, although there were 6 false negative results, specially for pancreatic or retroperitoneal lesions. Overall accuracy for abdominal procedures was 88%. It is concluded that CT guided fine needle biopsy is a safe and accurate method for the diagnosis of thoracic and abdominal lesions.

Abdominal Neoplasms↗
PubMed1991

Joubert syndrome. Respiratory failure requiring home mechanical ventilation.

A case of Joubert's Syndrome with respiratory insufficiency requiring mechanical ventilatory support is presented. The case illustrates the complex clinical, ethical and technical aspects in the management of patients with this syndrome and the feasibility of extending mechanical ventilatory support to the home setting. A caring, knowledgeable care team and a loving supportive family are required to properly continue treatment in the home in cases of this rare syndrome.

Abnormalities, Multiple↗
PubMed1990

Osteoporosis associated with neuroleptic treatment. A case report.

A 28-year-old woman developed osteoporosis following seven years of neuroleptic use. She presented with amenorrhea and profuse galactorrhea of four years' duration. Dual photon absorptionometry demonstrated reduced bone mineral density in the femur and spine. Serum calcium, phosphorus and alkaline phosphatase were normal. The patient was started on bromocriptine, and her bone density, serum prolactin, dehydroepiandrosterone sulfate, and free and total testosterone improved. No deterioration in her psychiatric condition occurred.

Adult↗
PubMed1989

Cocaine, pregnancy, and postpartum intracerebral hemorrhage.

An increase in cocaine abuse by pregnant women has been associated with a range of maternal/fetal cardiovascular complications. Intracerebral hemorrhage has been reported as a cocaine-related complication, but has not previously been associated with pregnancy. We report a case of cocaine-associated intracerebral hemorrhage in the postpartum period which complicated the differential diagnosis and management of hypertension and seizures.

Adult↗
PubMed1989

[Myoclonus as a manifestation of respiratory encephalopathy].

The triad of mental abnormalities, neurologic disorders and abnormal electroencephalogram is central to respiratory encephalopathy. The development of myoclonus has been considered as a sign heralding the development of generalized seizures and coma. A patient with chronic bronchitis is reported in whom myoclonus was the predominant symptom. Myoclonus was generalized, symmetrical and synchronic, and electroencephalographic correlation was shown, with diffuse polyspike-high potential wave discharges coincident with myoclonus. Remarkably, they promptly responded to low doses of diazepam, with disappearance of clinical neurologic and electrical findings. These have not subsequently relapsed although the patient is not receiving specific therapy. The risk of the administration of diazepam was weighted against other specific drugs (clonacepam, pyracetam) which are probably better tolerated. The interest in the quicker response of diazepam prevailed. We propose the inclusion of myoclonus secondary to respiratory encephalopathy in the classification of myoclonic disorders.

Brain Diseases↗
PubMed1988

Bone density and reproductive hormones in patients with neuroleptic-induced hyperprolactinemia.

To study the prevalence of osteoporosis and hyperandrogenism in neuroleptic-induced hyperprolactinemia, the authors evaluated 10 patients. Three were amenorrheic, while seven had oligomenorrhea. Nine patients had galactorrhea. The Ferriman-Gallway hirsutism score was 12 +/- 2. Vaginal smear maturation value was 53 +/- 8. Bone density, measured by dual photon absorptionometry in the spine, femoral neck, Ward's triangle, and trochanteric region, was 98 +/- 1.5, 92.7 +/- 3, 88.5 +/- 4.2, and 92.6 +/- 3.3 percentile of controls matched for age, sex, weight, and ethnicity, respectively. Serum prolactin, follicle-stimulating hormone (FSH), luteinizing hormone (LH), dehydroepiandrosterone sulfate (DHEAS), total testosterone, and free testosterone were 82 +/- 10 ng/ml, 8.5 +/- 1.1 mIU/ml, 11.1 +/- 2.6 mIU/ml, 4695 +/- 594 ng/ml, 90 +/- 17 ng%, and 2.36% +/- 0.3%, respectively. Serum thyroid-stimulating hormone (TSH) and free thyroxine index were normal. Bone density strongly correlated with vaginal maturation value (r = 0.904, P less than 0.01). It is concluded that (1) neuroleptic-induced hyperprolactinemia is associated with hirsutism and androgen excess primarily of adrenal origin and (2) a subset of these patients is at an increased risk of developing osteoporosis. It may be possible to identify patients at risk of osteoporosis by examining vaginal smears for maturation value. Early detection and management are imperative in this group of patients.

Adult↗