Atrial and brain natriuretic peptide in adrenal steroidogenesis.

We elucidated the role of atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) in human and bovine adrenocortical steroidogenesis. The urinary volume, sodium excretion and cyclic GMP (cGMP) excretion and plasma cGMP were markedly increased by the synthetic alpha-human ANP (alpha-hANP) infusion in healthy volunteers. Plasma arginine vasopressin (AVP) and aldosterone levels were significantly suppressed. Both ANP and BNP inhibited aldosterone, 19-OH-androstenedione, cortisol and DHEA secretion dose-dependently and increased the accumulation of intracellular cGMP in cultured human and bovine adrenal cells. alpha-hANP significantly suppressed P450scc-mRNA in cultured bovine adrenal cells stimulated by ACTH. Autoradiography and affinity labeling of [125I]hANP, and Scatchard plot demonstrated a specific ANP receptor in bovine and human adrenal glands. Purified ANP receptor from bovine adrenal glands identified two distinct types of ANP receptors, one is biologically active, the other is silent. A specific BNP receptor was also identified on the human and bovine adrenocortical cell membranes. The binding sites were displaced by unlabelled ANP as well as BNP. BNP showed an effect possibly via a receptor which may be shared with ANP. The mean basal plasma alpha-hANP level was 25 +/- 5 pg/ml in young men. We confirmed the presence of ANP and BNP in bovine and porcine adrenal medulla. Plasma or medullary ANP or BNP may directly modulate the adrenocortical steroidogenesis. We demonstrated that the lack of inhibitory effect of alpha-hANP on cultured aldosterone-producing adenoma (APA) cells was due to the decrease of ANP-specific receptor, which caused the loss of suppression of aldosterone and an increase in intracellular cGMP.     

Intracellular free Ca2+ concentration in fowl spermatozoa and its relationship to motility and respiration in spermatozoa.

The ability of fowl spermatozoa to accumulate and de-esterify the intracellular fluorescent Ca2+ indicator fura-2 was established. The cytosolic Ca2+ concentrations, measured by this technique, did not change after the addition of 1 mmol EGTA l-1. Subsequently, addition of the calcium ionophore A23187 caused a reduction in cytosolic Ca2+ concentrations, presumably by efflux of Ca2+ from the spermatozoa. Intracellular free Ca2+ concentrations were then significantly increased by the addition of 1 mmol CaCl2 l-1. The motility of demembranated spermatozoa gradually decreased after the addition of EGTA alone or EGTA with A23187, but was instantly restored by the addition of CaCl2 in the presence of both EGTA and A23187. Unlike demembranated spermatozoa, intact spermatozoa maintained their motility, even after the addition of EGTA, but their motility was reduced by the addition of A23187 in the presence of EGTA. The addition of A23187 also reduced the rate of oxygen consumption, but not the ATP concentrations in intact spermatozoa. These results demonstrate that the motility and respiration of fowl spermatozoa are strongly influenced by their intracellular Ca2+ concentrations.     

Observations on the effect of visual and olfactory ablation on the swimming behavior of migrating adult chum salmon,Oncorhynchus keta

The behavior of chum slamon,Oncorhynchus keta, was studied using ultrasonic telemetry in the waters off the Okhotsk coast of Hokkaido from 1979 to 1981. Thirty-six adult fish were outfitted with a 50 KHz ultrasonic transmitter provided with either a depth sensor, depth/illumination sensors, or depth/ temperature sensors. Twenty-one of the experimental fish were used as controls and left intact. Of the remaining fish, six had their sight destroyed, seven had their olfactory nerves severed or their nares filled with wax, one had both senses destroyed, and one had a sham operation, but had only a shallow transverse cut made in the skin over the olfactory nerves. Control fish and the fish with the sham operation initially swam in a horizontal zigzag pattern while fish with an obliterated sense swam in a similar pattern but to a reduced extent. The horizontal and vertical speeds of the intact fish were faster than those of the fish with a destroyed sense. Amplitude of vertical movements of the intact fish stretched from surface to bottom, while fish with destroyed vision tended towards midwater. Fish without an olfactory sense tended to be at the surface or near the bottom. Regular vertical movement would be effective in refreshing olfactory epithelia which might become acclimated when exposed to one odor. The vertical movements can be found in the horizontal zigzag movements in coastal and near shore migrations, as well as in the stream phase of homing. The zigzag movements, both horizontal and vertical, allow the fish to sense the water masses and locate the correct tributary.     

Schistosoma japonicum: Immediate hypersensitivity reactions to egg antigen in humans and rodents

Immediate hypersensitivity reactions in schistosoma japonicum infections were examined in both man and experimental animals. In man higher reaction to soluble egg antigen than to adult worm antigen was detected by the use of the radioallergosorbent test (RAST). Blood-collecting filter paper can be used in RAST for seroepidemiological study in place of a skin test. Reaginic antibody formation against egg antigen was detected at the approximate time of egg deposition in strains of mice, Mongolian gerbils, cotton rats, and laboratory rats by the use of passive cutaneous anaphylaxis or Prausnitz-Küstner-type skin tests. At the same time circumoval precipitin tests were positive. Results with athymic nude mice suggest that these reactions are T-cell dependent. No detectable reagin synthesis against adult worm antigen was found in the animals so far examined, confirming stronger allergenic reaction to egg antigen than to that of adult worms in S. japonicum infections in man and animals.     

Binding of HIV-1 gp120 to DC-SIGN Promotes ASK-1-Dependent Activation-Induced Apoptosis of Human Dendritic Cells

During disease progression to AIDS, HIV-1 infected individuals become increasingly immunosuppressed and susceptible to opportunistic infections. It has also been demonstrated that multiple subsets of dendritic cells (DC), including DC-SIGN(+) cells, become significantly depleted in the blood and lymphoid tissues of AIDS patients, which may contribute to the failure in initiating effective host immune responses. The mechanism for DC depletion, however, is unclear. It is also known that vast quantities of viral envelope protein gp120 are shed from maturing HIV-1 virions and form circulating immune complexes in the serum of HIV-1-infected individuals, but the pathological role of gp120 in HIV-1 pathogenesis remains elusive. Here we describe a previously unrecognized mechanism of DC death in chronic HIV-1 infection, in which ligation of DC-SIGN by gp120 sensitizes DC to undergo accelerated apoptosis in response to a variety of activation stimuli. The cultured monocyte-derived DC and also freshly-isolated DC-SIGN(+) blood DC that were exposed to either cross-linked recombinant gp120 or immune-complex gp120 in HIV(+) serum underwent considerable apoptosis after CD40 ligation or exposure to bacterial lipopolysaccharide (LPS) or pro-inflammatory cytokines such as TNFα and IL-1β. Furthermore, circulating DC-SIGN(+) DC that were isolated directly from HIV-1(+) individuals had actually been pre-sensitized by serum gp120 for activation-induced exorbitant apoptosis. In all cases the DC apoptosis was substantially inhibited by DC-SIGN blockade. Finally, we showed that accelerated DC apoptosis was a direct consequence of excessive activation of the pro-apoptotic molecule ASK-1 and transfection of siRNA against ASK-1 significantly prevented the activation-induced excessive DC death. Our study discloses a previously unknown mechanism of immune modulation by envelope protein gp120, provides new insights into HIV immunopathogenesis, and suggests potential therapeutic approaches to prevent DC depletion in chronic HIV infection.     

Determination of the Role of DDX3 a Factor Involved in Mammalian RNAi Pathway Using an shRNA-Expression Library

RNA interference (RNAi) is an endogenous RNA-destruction phenomenon induced by certain double-stranded RNAs (dsRNAs). In RNAi, dsRNAs are processed into small interfering RNAs (siRNAs) which in turn trigger the cleavage of the target mRNA. Here, using a short hairpin RNA-expression library, we identified a DEAD-box helicase 3, DDX3, as an essential factor involved in RNAi pathway and revealed that DDX3 is colocalized with Ago2, an essential factor in RNAi pathway that cleaves target mRNA. Results of experiments with a dominant negative mutant of DDX3 further confirmed that this factor affects the RNAi activity. Together, DDX3 functions to assure mammalian RNAi pathway. Together, our results indicate that DDX3 is a new key molecule to understand the molecular mechanism underlying RNAi pathway in mammals.     

Insulin resistance dysregulates CYP7B1 leading to oxysterol accumulation: a pathway for NAFL to NASH transition

NAFLD is an important public health issue closely associated with the pervasive epidemics of diabetes and obesity. Yet, despite NAFLD being among the most common of chronic liver diseases, the biological factors responsible for its transition from benign nonalcoholic fatty liver (NAFL) to NASH remain unclear. This lack of knowledge leads to a decreased ability to find relevant animal models, predict disease progression, or develop clinical treatments. In the current study, we used multiple mouse models of NAFLD, human correlation data, and selective gene overexpression of steroidogenic acute regulatory protein (StarD1) in mice to elucidate a plausible mechanistic pathway for promoting the transition from NAFL to NASH. We show that oxysterol 7α-hydroxylase (CYP7B1) controls the levels of intracellular regulatory oxysterols generated by the “acidic/alternative” pathway of cholesterol metabolism. Specifically, we report data showing that an inability to upregulate CYP7B1, in the setting of insulin resistance, results in the accumulation of toxic intracellular cholesterol metabolites that promote inflammation and hepatocyte injury. This metabolic pathway, initiated and exacerbated by insulin resistance, offers insight into approaches for the treatment of NAFLD.     

Unique active site formation in a novel galactose 1-phosphate uridylyltransferase from the hyperthermophilic archaeon Pyrobaculum aerophilum

A gene encoding galactose 1-phosphate uridylyltransferase (GalT) was identified in the hyperthermophilic archaeon Pyrobaculum aerophilum. The gene was overexpressed in Escherichia coli, after which its product was purified and characterized. The expressed enzyme was highly thermostable and retained about 90% of its activity after incubation for 10 minutes at temperatures up to 90°C. Two different crystal structures of P. aerophilum GalT were determined: the substrate-free enzyme at 2.33 Å and the UDP-bound H140F mutant enzyme at 1.78 Å. The main-chain coordinates of the P. aerophilum GalT monomer were similar to those in the structures of the E. coli and human GalTs, as was the dimeric arrangement. However, there was a striking topological difference between P. aerophilum GalT and the other two enzymes. In the E. coli and human enzymes, the N-terminal chain extends from one subunit into the other and forms part of the substrate-binding pocket in the neighboring subunit. By contrast, the N-terminal chain in P. aerophilum GalT extends to the substrate-binding site in the same subunit. Amino acid sequence alignment showed that a shorter surface loop in the N-terminal region contributes to the unique topology of P. aerophilum GalT. Structural comparison of the substrate-free enzyme with UDP-bound H140F suggests that binding of the glucose moiety of the substrate, but not the UDP moiety, gives rise to a large structural change around the active site. This may in turn provide an appropriate environment for the enzyme reaction.     

The Spherical Expansion of H2 Dimer Interaction Energy by Double Symmetry-Adapted Perturbation Theory

Abstract

The weak interaction energy of H₂ dimer is studied by double symmetry-adapted perturbation theory (SAPT) within second-order of intermolecular and intramonomer perturbation for molecular simulations. The assumed orientations of H₂ dimer are linear, parallel, T type and X type. Among four orientations T orientation is the most stable, while linear orientation is the most repulsive. The second-order dispersion energy E _disp ⁽²⁾ is the most attractive contribution in all orientations. The interaction energy has the anisotropy, so we expressed our total interaction energy by the spherical expansion to compare with the experimental value. The isotropic interaction energy is about 85% of the experimental value.