Effect of partial proteolysis on the activation energy of beta-n-acetylhexosaminidase precursor and mature forms.

Using 3,3'-diclorophenolsulfoftaleinil N-acetyl-beta-D-glucosaminide as a substrate, the apparent activation energy of beta-N-acetylhexosaminidase (Hex) was determined in samples of plasma and urine, as well as in leukocyte and platelet lysates. Incubation with papain produced an increase in this thermodynamic variable for plasma Hex (precursor forms with high molecular mass) that would be caused by the proteolytic action of papain on the Hex A isoenzyme. However, digestion with papain did not significantly modify the activation energy of Hex in leukocyte and platelet lysates (mature enzymatic forms). In 11 healthy subjects and 28 patients with different renal diseases, no statistically significant differences were found with regard to the values obtained in cellular lysates for variations in the activation energy of urinary Hex, regardless of whether they presented normoalbuminuria, microalbuminuria or macroalbuminuria. These results support the hypothesis that even in patients with proteinuria, no significant amounts of plasma Hex precursor forms are found in urine samples, and the source of the enzyme activity is the kidney itself.     

Hormonal and Metabolic Disorders of Human Immunodeficiency Virus Infection and Substance Abuse

There are an estimated 200 million users of an illicit drug in the world today. In addition, an estimated 40 million people are infected with the human immunodeficiency virus (HIV) and an estimated 180 million people are infected with the hepatitis C virus (HCV). Both the use of an illicit drug and the co-occurrence of infections are associated with a multitude of medical and health consequences including hormonal and metabolic disorders. Thus, the National Institute on Drug Abuse (NIDA), a part of the National Institutes of Health (NIH) hosted a workshop on hormonal and metabolic disorders of HIV among substance abusers. A number of clinicians and scientists participated and discussed a wide range of issues concerning hormones, nutrition and metabolic complications in HIV and substance abuse. Their observations and the recommendations they made for future research are presented in these proceedings. The readers are encouraged to contact the NIH staff (JK, FV) for technical guidance and programmatic priorities on the subject and directly contact the individual authors for collaborations.     

Complete maternal uniparental isodisomy of chromosome 4 in a subject with major depressive disorder detected by high density SNP genotyping arrays.

Uniparental isodisomy (iUPD) is a rare genetic condition caused by non-disjunction during meiosis that ultimately leads to a duplication of either the maternal or paternal chromosome in the affected individual. Two types of disorders can result, those due to imprinted genes and those due to homozygosity of recessive disease-causing mutations. Here, we describe the third known case of complete chromosome 4 iUPD of maternal origin. This condition became apparent during whole genome linkage studies of psychiatric disorders in the Portuguese population. The proband is an adult female with normal fertility and no major medical complaints, but a history of major depressive disorder and multiple suicide attempts. The proband's siblings and parents had normal chromosome 4 genotypes and no history of mood disturbance. A brief review of other studies lends support for the possibility that genes on chromosome 4 might confer risk for mood disorders. We conclude that chromosome 4 maternal uniparental disomy (UPD) is a rare disorder that may present with a major depressive phenotype. The lack of a common disease phenotype between this and two other cases of chromosome 4 iUPD [Lindenbaum et al. [1991] Am J Med Genet 49(Suppl 285):1582; Spena et al. [2004] Eur J Hum Genet 12:891-898) would suggest that there is no vital maternal gene imprinting on chromosome 4. However, since there is no reported case of paternal chromosome 4 UPD, paternal gene imprinting on chromosome 4 cannot be excluded.     

Left pulmonary artery evaluation through transesophageal echocardiography.

INTRODUCTION: Transesophageal echocardiography (TEE) has become increasingly useful in the study of patients with suspected pulmonary thromboembolism. OBJECTIVE: The aim of this study was to prospectively evaluate the usefulness of TEE in the study of the distal part of the left pulmonary artery (LPA) as well as the influence of this procedure on total echocardiographic exam duration. METHODOLOGY: A prospective study in two groups of consecutive patients referred for TEE with a one- year interval between evaluation of Group A: 33 patients, 17 male, mean age 54 +/- 24 years, and Group B: 42 patients, 20 male, mean age 48 +/- 27 years (p = NS). The procedure was considered long when it took more than 3 min to evaluate the distal part of the LPA. RESULTS: In group A we were able to visualize the distal part of the LPA in 24 patients (73%) without significant prolongation of total exam duration in 16 patients (48% of group A). In one of the patients with suspected pulmonary thromboembolism thrombi were only observed in the distal part of the LPA. In group B we were able to visualize the distal part of the LPA in 36 patients (86%) without significant prolongation of total exam duration in 26 patients (61% of group B). CONCLUSIONS: 1. Visualization of the distal part of the LPA was possible in more patients, and with TEE time prolongation in less patients, in group B. These differences can be accounted for by the training of the operator in this technique. 2. The importance of visualization of this part of the LPA in guiding treatment in the subset of patients with pulmonary thromboembolism confirms the usefulness of this specific procedure.     

Direct effect of acute metabolic and respiratory acidosis on parathyroid hormone secretion in the dog.

Because both metabolic (Met Acid) and respiratory acidosis (Resp Acid) have diverse effects on mineral metabolism, it has been difficult to establish whether acidosis directly affects parathyroid hormone (PTH) secretion. Our goal was to determine whether acute Met Acid and Resp Acid directly affected PTH secretion. Three groups of dogs were studied: control, acute Met Acid induced by HCl infusion, and acute Resp Acid induced by hypoventilation. EDTA was infused to prevent acidosis-induced increases in ionized calcium, but more EDTA was needed in Met Acid than in Resp Acid. The PTH response to EDTA-induced hypocalcemia was evaluated also. Magnesium needed to be infused in groups receiving EDTA to prevent hypomagnesemia. The half-life of intact PTH (iPTH) was determined during hypocalcemia when PTH was measured after parathyroidectomy. During normocalcemia, PTH values were greater (p < 0.05) in Met Acid (92 +/- 19 pg/ml) and Resp Acid (77 +/- 22 pg/ml) than in controls (27 +/- 5 pg/ml); the respective pH values were 7.23 +/- 0.01, 7.24 +/- 0.01, and 7.39 +/- 0.02. The maximal PTH response to hypocalcemia was greater (p < 0.05) in Met Acid (443 +/- 54 pg/ml) than in Resp Acid (267 +/- 37 pg/ml) and controls (262 +/- 48 pg/ml). The half-life of PTH was greater (p < 0.05) in Met Acid than in controls, but the PTH secretion rate also was greater (p < 0.05) in Met Acid than in the other two groups. In conclusion, (1) both acute Met Acid and Resp Acid increase PTH secretion when the ionized calcium concentration is normal; (2) acute Met Acid may increase the bone efflux of calcium more than Resp Acid; (3) acute Met Acid acts as a secretogogue for PTH secretion because it enhances the maximal PTH response to hypocalcemia.     

P2X(7) receptors are expressed during mouse nephrogenesis and in collecting duct cysts of the cpk/cpk mouse.

BACKGROUND: Purinergic receptors are cell-surface molecules that bind extracellular nucleotides, notably ATP. The P2X family includes seven nonselective ion channels with one member, P2X(7), implicated in cytolytic pore formation and cell death. MATERIALS AND METHODS: We sought P2X(7) expression in mouse nephrogenesis and cpk/cpk renal cyst growth, conditions in which both proliferation and apoptosis are prominent. RESULTS: P2X(7) immunolocalized to condensed metanephric mesenchyme: both proliferation and apoptosis were detected in this compartment, assessed by proliferating cell nuclear antigen expression and propidium iodide-stained pyknotic nuclei respectively. Later in nephrogenesis, P2X(7) was detected in collecting ducts, a pattern persisting to maturity. A mesenchymal to epithelial shift of P2X(7) expression was also documented in ureter development. In cpk/cpk kidneys, P2X(7)-expressing collecting duct cysts dominated histology from two weeks until four weeks after birth, when animals die from uremia. In polycystic kidneys pyknotic nuclei were rarely identified in P2X(7)-expressing epithelia, but were detected between cysts, consistent with a non-apoptotic role for P2X(7) in cyst enlargement. CONCLUSION: P2X(7) is expressed during normal nephrogenesis and in a model of congenital polycystic kidney disease. Further experiments are necessary to define possible functions of P2X(7) in these settings.     

Differential Effects of Anesthetics on Mitochondrial KATP Channel Activity and Cardiomyocyte Protection

Background: Mitochondrial adenosine triphosphate-sensitive potassium (mitoKATP) channels play a pivotal role in mediating cardiac preconditioning. The effects of intravenous anesthetics on this protective channel have not been investigated so far, but would be of importance with respect to experimental as well as clinical medicine.

Methods: Live cell microscopy was used to visualize and measure autofluorescence of flavoproteins, a direct reporter of mitoKATP channel activity, in response to the direct and highly selective mitoKATP channel opener diazoxide, or to diazoxide following exposure to various anesthetics commonly used in experimental and clinical medicine. A cellular model of ischemia with subsequent hypoosmolar trypan blue staining served to substantiate the effects of the anesthetics on mitoKATP channels with respect to myocyte viability.

Results: Diazoxide-induced mitoKATP channel opening was significantly inhibited by the anesthetics R-ketamine, and the barbiturates thiopental and pentobarbital. Conversely, urethane, 2,2,2-trichloroethanol (main metabolite of [alpha]-chloralose and chloral hydrate), and the opioid fentanyl potentiated the channel-opening effect of diazoxide, which was abrogated by coadministration of chelerythrine, a specific protein kinase C inhibitor. S-ketamine, propofol, xylazine, midazolam, and etomidate did not affect mitoKATP channel activity. The significance of these modulatory effects of the anesthetics on mitoKATP channel activity was substantiated in a cellular model of simulated ischemia, where diazoxide-induced cell protection was mitigated by R-ketamine and the barbiturates, while urethane, 2,2,2-trichloroethanol, and fentanyl potentiated myocyte protection.     

Thyroid Hormone in the Treatment of Post-transplant Acute Tubular Necrosis (ATN)

Delayed graft function (DGF) in cadaver kidney transplants is a common problem and is often due to acute tubular necrosis (ATN). DGF in transplants may have a deleterious effect on long-term graft survival. Since thyroid hormone has been shown to hasten recovery from ATN in experimental models, we designed a trial to determine if a defined course of triiodothyronine (T3) would improve the short- or long-term outcome of patients with DGF in cadaveric transplants. A prospective, randomized, placebo controlled, double blind trial of T3 was carried out in patients with DGF in cadaveric renal transplants. End-points were percentage requiring dialysis, percentage recovering function, time to recovery and length of hospital stay. Long-term outcomes were percentage grafts functioning at 1 year and mean serum creatinine at 1 year. Forty-four patients were randomized to receive either T3 or placebo. Three patients were dropped from each group when early biopsies disclosed that DGF was due to rejection. The groups were well matched by age, cold ischemia time of the graft, and percentage reactivity to a random panel of antigens. Baseline thyroid function studies, including T3, reverse T3 (rT3), and thyroid stimulating hormone (TSH) levels, were similar between the two groups and typical of 'euthyroid-sick syndrome'. T3 had no effect on percentage requiring dialysis, time to recovery, percentage recovering function, or length of stay. At 1 year follow-up, graft function was similar in both groups and significantly lower than that seen in patients with good initial function. Thyroid hormone, given early in the course of DGF in cadaver kidney recipients, had no effect on the course of DGF. Long-term graft function is impaired in patients who experience post-transplant DGF compared to those who have good initial function.     

Comparison of yogurt, soybean, casein, and amino acid–based diets in children with persistent diarrhea

Although previous studies have shown successful treatment of persistent diarrhea (PD) with the use of yogurt-based diets, some recent ones speculate the need of special formulas for the nutritional management of PD complicated cases. In the present study, we tested the hypothesis that the consumption of 3 lactose-free diets, with different degrees of complexity, is associated with lower stool output and shorter duration of diarrhea when compared with the use of a yogurt-based one on the nutritional management of PD. A total of 154 male infants, aged between 1 and 30 months, with PD and with or without dehydration, were randomly assigned to 1 of 4 treatment groups. Throughout the study, the patients were placed in a metabolic unit; their body weights and intakes of oral rehydration solution, water, and formula diets, in addition to outputs of stool, urine, and vomit, were measured and recorded at 24-hour intervals. Four different diets were used in this study: diet 1, yogurt-based formula; diet 2, soy-based formula; diet 3, hydrolyzed protein-based formula; and diet 4, amino acid–based formula. Throughout the study, only these formula diets were fed to the children. The data showed that children fed the yogurt-based diet (diet 1) or the amino acid–based diet (diet 4) had a significant reduction in stool output and in the duration of diarrhea. The use of an inexpensive and worldwide-available yogurt-based diet is recommended as the first choice for the nutritional management of mild to moderate PD. For the few complicated PD cases, when available, a more complex amino acid–based diet should be reserved for the nutritional management of these unresponsive and severe presentations. Soy-based or casein-based diets do not offer any specific advantage or benefits and do not seem to have a place in the management of PD.