Chronic exposure of rats to hypoxic environment alters the mechanism of energy transfer in myocardium

We have investigated the effect of chronic exposure of rats to an hypoxic environment (10% O2; 3 weeks), on the first step of the intracellular energy transfer process in the myocardium, i.e. the transfer at mitochondrial level of high energy bonds from ATP to creatine. In the left ventricles from rats adapted to normobaric hypoxia, we observed, using the permeabilized fiber technique, that the stimulatory effect of creatine on the mitochondrial respiration in presence of a low ADP concentration (0.1 mM) was attenuated when compared to control. Furthermore, the creatine-induced decrease of the apparent K(m) for ADP of the mitochondrial respiration, which is observed in control, was significantly reduced. Both the basal and maximal respiratory rates of the fibers were unchanged by the hypoxic exposure of the rats. A significant decrease of the total creatine kinase activity from 755 to 630 IU/g wet weight (for control and hypoxic rats, respectively) was detected and was accompanied by a 25% decrease in mitochondrial isoform activity (mitoCK) and in the mitoCK/citrate synthase ratio. In the right ventricles, identical alterations in the effect of creatine on apparent K(m) for ADP were observed while we did not detect any changes in CK activity. The decrease in mitoCK activity and the fall in the reactivity of respiration to creatine could be interpreted as a mechanism for downregulating oxygen demand during chronic hypoxia. The consequences of such alterations on energy metabolism of cardiomyocytes under conditions of reduced oxygen supply are discussed.     

Triplex formation at physiological pH by oligonucleotides incorporating 5-Me-dC-(N4-spermine)

Oligonucleotide (ODN) directed triplex formation has therapeutic importance and depends on Hoogsteen hydrogen bonds between a duplex DNA and a third strand. While T*A:T triplets are formed at neutral pH, C+*G:C are favoured at acidic pH. Herein it is shown that 18-mer ODN containing spermine conjugated to 5-Me-dC at N4 (1-5), form triplexes with complementary 24-mer duplex 8:9 at neutral pH (7.3, 100 mM NaCl). Under such conditions, control ODN's carrying dC (6) or 5-Me-dC (7) did not show any triple helix formation. Remarkably, the triplexes from spermine-conjugates (1-5) have foremost stability at neutral pH (7.1), unlike the behavior of normal ODN's where optimal stability is at acidic pH (5.5). These results have importance in designing oligonucleotides for antigene applications.     

Prevention of T cell anergy by signaling through the gamma c chain of the IL-2 receptor

When stimulated through their antigen receptor without requisite costimulation, T cells enter a state of antigen-specific unresponsiveness termed anergy. In this study, signaling through the common gamma chain of the interleukin-2 (IL-2), IL-4, and IL-7 receptors in the presence of antigen was found to be sufficient to prevent the induction of anergy. After culture with IL-2, IL-4, or IL-7, Jak3 kinase was tyrosine-phosphorylated, which correlated with the prevention of anergy. Therefore, a signal through the common gamma chain may regulate the decision of T cells to either clonally expand or enter a state of anergy.     

Choline derived from the phosphatidylcholine specific phospholipase D is not directly available for the CDP choline pathway in phorbol ester-treated C3H10T1/2 Cl 8 fibroblasts

To clarify the mechanism of capsaicin-induced primary neuronal cell death, newborn and adult rats were given a subcutaneous injection of capsaicin (50 mg/kg). Neonatal capsaicin injection induced neuronal apoptosis in the trigeminal ganglion. Apoptotic neurons had peripheral stacks of long parallel endoplasmic reticulum that are characteristic to primary neurons of the B-type, and exhibited nucleoplasmic condensation, nuclear shrinkage and cytoplasmic fragmentation. Light microscopically, apoptotic neurons exhibited a sign of DNA fragmentation as revealed by a nick end labelling method. The proportion of apoptotic cells was quite low during the first 12 h after capsaicin injection (<1%), rapidly increase to 10.44% by 24 h, and decreased to 0.29% by 48 h. Normal and vehicle control levels of apoptosis were <1%. Nerve growth factor (NGF, 0.5 mg/kg) simultaneously administered with capsaicin reduced the incidence of apoptosis by about 35% at 24 h post-injection. Neonatal transection of the infraorbital nerve induced neuronal apoptosis similar to that produced by the neonatal capsaicin in the maxillary division of the trigeminal ganglion. Unlike capsaicin, however, the neurotomy-induced apoptosis was seen in neurons of both the A- and B-types. Neither the capsaicin injection nor the neurotomy induced apoptosis in adult rats, though mitochondrial swelling similar to that seen at 0.5 h after neonatal capsaicin was observed after capsaicin injection in adults. The results indicate that the capsaicin-induced and nerve injury-induced primary neuronal damages in newborn rats share a common final pathway, apoptosis.     

Epidemiology of giardiasis and cryptosporidiosis in Jamaica

We report the findings of a cross-sectional epidemiologic study of Giardia lamblia and Cryptosporidum infections in Jamaica. Three hundred twenty eight stool samples from patients less than one to 81 years of age were examined using formalin-ether concentration for G. lamblia, Zeihl-Neelsen staining for Cryptosporidium, and the Prospect rapid enzyme immunoassay (EIA; Alexon, Sunnyvale, CA) for parasite diagnosis. The Prospect Giardia rapid assay detected 17 cases of G. lamblia infection compared with six by formalin-ether concentration. However, the Prospect Cryptosporidum EIA did not increase the rate of detection of Cryptosporidum when compared with Zeihl-Neelsen staining. Cryptosporidum infections were most frequently diagnosed in children less than five years old and prevalence decreased with age. In contrast, the prevalence of giardiasis increased as children became older. There were no associations between the infections and stool consistency, clinical manifestations, or sex of the individuals. The contribution of the parasites to childhood morbidity will depend on accurate laboratory diagnosis.     

Therapeutic effects of prolonged normovolemic hemodilution in patients with Raynaud's disease

51 patients with Raynaud's disease has undergone prolonged normovolemic hemodilution (PNH) with a good effect. Disappearance of painful syndrome in 78.43% of cases, and a significant decrease of pains in 21.37% of cases was demonstrated. Complete healing of ulcero-necrotic defects of end phalanges of the hands was observed in 7 patients of 10 and partial--in 3. A significant remission of the disease in follow-up period was obtained. Thus, prolonged normovolemic hemodilution has resulted in significant improvement of short- and long-term results of treatment in patients with Raynaud's disease.     

Influence of cell polarity on retrovirus-mediated gene transfer to differentiated human airway epithelia

Gene transfer with recombinant murine leukemia viruses (MuLV) provides the potential to permanently correct inherited lung diseases, such as cystic fibrosis (CF). Several problems prevent the application of MuLV-based recombinant retroviruses to lung gene therapy: (i) the lack of cell proliferation in mature pulmonary epithelia, (ii) inefficient gene transfer with a vector applied to the apical surface, and (iii) low titers of many retroviral preparations. We found that keratinocyte growth factor (KGF) stimulated proliferation of differentiated human tracheal and bronchial epithelia. Approximately 50% of epithelia divided in response to KGF as assessed by bromodeoxyuridine histochemistry. In airway epithelia stimulated to divide with KGF, high-titer ampho- and xenotropic enveloped vectors preferentially infected cells from the basal side. However, treatment with hypotonic shock or EGTA transiently increased transepithelial permeability, enhancing gene transfer with the vector applied to the mucosal surfaces of KGF-stimulated epithelia. Up to 35% of cells expressed the transgene after gene transfer. By using this approach, cells throughout the epithelial sheet, including basal cells, were targeted. Moreover, the Cl- transport defect in differentiated CF airway epithelia was corrected. These findings suggest that barriers to apical infection with MuLV can be overcome.     

Polyclonal antibodies from blood and cerebrospinal fluid of patients with multiple sclerosis effectively hydrolyze DNA and RNA

It is known that in the blood of patients with some autoimmune diseases catalytically active antibodies hydrolyzing proteins, DNA, and RNA may be detected. In the present work homogeneous preparations of IgG antibodies (Ab) possessing high affinity for nucleic acids (NA) were obtained for the first time from blood and cerebrospinal fluid of patients with multiple sclerosis (MS). The fraction of IgG Ab as well as its Fab fragments and isolated light chains of both kappa- and lambda-types were shown to catalyze effectively the hydrolysis of DNA and RNA. It is shown by different methods that the capability for nucleic acid hydrolysis is an intrinsic property of the polyclonal Ab. NA-hydrolyzing Ab were detected in the blood of 69 of 72 and in the cerebrospinal fluid of 5 of 5 examined MS patients, while they were not detected in the blood of any of 50 healthy donors examined. Comparison of relative rates of RNA hydrolysis and of the substrate specificity in hydrolysis of various model RNAs--cCMP, poly(U), poly(A), and poly(C)--revealed pronounced differences of MS antibodies from ribonucleases of human blood, ribonuclease A, and all earlier described abzymes. The abzymes are usually characterized by relatively low specific activities in comparison with that of normal enzymes catalyzing analogous reactions. Ab from the blood of MS patients are the first example of autoabzymes whose specific activity in RNA hydrolysis is comparable or even higher than that of pancreatic ribonuclease A--one of the most active RNA-hydrolyzing enzymes.     

Dyslipidemias and autoimmune diseases

The paper summarizes the results of over 30-year studies dealt with dyslipidemias and autoimmune diseases. The teaching of the antiphospholipid syndrome (APS) has aroused interest in the problem. The experience gained shows changes in the blood cholesterol transport system. Patients with systemic lupus erythematosus (SLE) have higher levels of low density lipoprotein cholesterol and lower concentrations of high density lipoprotein (HDL) cholesterol, apolipoprotein A than the controls. The quantitative and qualitative changes in particles result in decreased acceptance of cholesterol from the membrane of a cell and tissues, which promotes the development of vascular diseases. Lipoprotein (a) may be an additional risk factor for thrombosis chiefly of coronary arteries, in patients with SLE and APS. Increased levels of oxidized low density proteins having atherogenic activity were found mainly in patients with SLE. The use of corticosteroids causes the changes in the spectrum of blood lipids, which together with other factors (thrombosis, vasculopathy, thrombocytopenia, etc.) create good conditions for the development of atherosclerosis, which determines the necessity of correcting the parameters of blood lipid transport not only to prevent vascular disorders but to improve the general life prognosis in SLE patients.     

Effect of alcohol consumption on adult and aged bone: composition, morphology, and hormone levels of a rat animal model

To determine the effect of life-long alcohol consumption on the adult and aged rat model, 4-week-old, female Sprague-Dawley rats were divided into three diet groups. Alcohol-treated animals were fed a modified Lieber-DeCarli diet ad libitum containing 35% ethanol-derived calories, whereas the pair-fed animals (weight-matched to ethanol rats) received an isocaloric liquid diet in which maltose-dextrin substituted calories supplied by ethanol. Chow animals were fed a standard rat chow ad libitum. Proximal tibiae (primarily cancellous bone) and femora (primarily cortical bone) were removed for analysis after 3, 6, 9, 12, or 18 months on the diets. Serum was collected for analysis of calcium levels, the calcium regulating hormones; parathyroid hormone, 25-hydroxyvitamin D, calcitonin, corticosterone, estradiol, testosterone, and IGF-1. Creatinine, SGOT/AST, and SGPT/ALT levels were measured to determine kidney and liver integrity. Previous studies, with young animals, showed that chronic alcohol consumption during the age of bone development reduced bone density and bone mass in both cortical and cancellous bone. The present study demonstrates that these reductions last throughout life, whereas morphological values, such as length and diameter, attain control levels. Calcium regulating hormones and sex hormones are essentially normal and do not appear to be the primary causative agent for adult alcohol-induced osteopenia, but it appears to be due to a more direct effect of alcohol on bone cells.