Hypothyroidism provides resistance to kidney mitochondria against the injury induced by renal ischemia-reperfusion

Massive Ca(2+) accumulation in mitochondria, plus the stimulating effect of an inducing agent, i.e., oxidative stress, induces the so-called permeability transition, which is characterized by the opening of a nonspecific pore. This work was aimed at studying the influence of thyroid hormone on the opening of such a nonspecific pore in kidney mitochondria, as induced by an oxidative stress. To meet this objective, membrane permeability transition was examined in mitochondria isolated from kidney of euthyroid and hypothyroid rats, after a period of ischemia/reperfusion. It was found that mitochondria from hypothyroid rats were able to retain accumulated Ca(2+) to sustain a transmembrane potential after Ca(2+) addition, as well as to maintain matrix NAD(+) and membrane cytochrome c content. The protective effect of hypothyroidism was clearly opposed to that occurring in ischemic reperfused mitochondria from euthyroid rats. Our findings demonstrate that these mitochondria were unable to preserve selective membrane permeability, except when cyclosporin A was added. It is proposed that the protection is conferred by the low content of cardiolipin found in the inner membrane. This phospholipid is required to switch adenine nucleotide translocase from specific carrier to a non-specific pore.     

Genome-Wide Association Studies in an Isolated Founder Population from the Pacific Island of Kosrae

It has been argued that the limited genetic diversity and reduced allelic heterogeneity observed in isolated founder populations facilitates discovery of loci contributing to both Mendelian and complex disease. A strong founder effect, severe isolation, and substantial inbreeding have dramatically reduced genetic diversity in natives from the island of Kosrae, Federated States of Micronesia, who exhibit a high prevalence of obesity and other metabolic disorders. We hypothesized that genetic drift and possibly natural selection on Kosrae might have increased the frequency of previously rare genetic variants with relatively large effects, making these alleles readily detectable in genome-wide association analysis. However, mapping in large, inbred cohorts introduces analytic challenges, as extensive relatedness between subjects violates the assumptions of independence upon which traditional association test statistics are based. We performed genome-wide association analysis for 15 quantitative traits in 2,906 members of the Kosrae population, using novel approaches to manage the extreme relatedness in the sample. As positive controls, we observe association to known loci for plasma cholesterol, triglycerides, and C-reactive protein and to a compelling candidate loci for thyroid stimulating hormone and fasting plasma glucose. We show that our study is well powered to detect common alleles explaining ≥5% phenotypic variance. However, no such large effects were observed with genome-wide significance, arguing that even in such a severely inbred population, common alleles typically have modest effects. Finally, we show that a majority of common variants discovered in Caucasians have indistinguishable effect sizes on Kosrae, despite the major differences in population genetics and environment.     

Palmitoylation of inducible nitric-oxide synthase at Cys-3 is required for proper intracellular traffic and nitric oxide synthesis

A number of cell types express inducible nitric-oxide synthase (NOS2) in response to exogenous insults such as bacterial lipopolysaccharide or proinflammatory cytokines. Although it has been known for some time that the N-terminal end of NOS2 suffers a post-translational modification, its exact identification has remained elusive. Using radioactive fatty acids, we show herein that NOS2 becomes thioacylated at Cys-3 with palmitic acid. Site-directed mutagenesis of this single residue results in the absence of the radiolabel incorporation. Acylation of NOS2 is completely indispensable for intracellular sorting and .NO synthesis. In fact, a C3S mutant of NOS2 is completely inactive and accumulates to intracellular membranes that almost totally co-localize with the Golgi marker beta-cop. Likewise, low concentrations of the palmitoylation blocking agents 2-Br-palmitate or 8-Br-palmitate severely affected the .NO synthesis of both NOS2 induced in muscular myotubes and transfected NOS2. However, unlike endothelial NOS, palmitoylation of inducible NOS is not involved in its targeting to caveolae. We have created 16 NOS2-GFP chimeras to inspect the effect of the neighboring residues of Cys-3 on the degree of palmitoylation. In this regard, the hydrophobic residue Pro-4 and the basic residue Lys-6 seem to be indispensable for palmitoylation. In addition, agents that block the endoplasmic reticulum to Golgi transit such as brefeldin A and monensin drastically reduced NOS2 activity leading to its accumulation in perinuclear areas. In summary, palmitoylation of NOS2 at Cys-3 is required for both its activity and proper intracellular localization.     

Substituted aminobenzimidazole pyrimidines as cyclin-dependent kinase inhibitors

A series of aminobenzimidazole-substituted pyrimidines were synthesized and evaluated for biochemical activity against CDK1. A high-speed parallel synthesis approach enabled the identification of a potent lead series having improved potency in the CDK1 assay (IC(50)<10nM). Cell cycle analysis showed that the compounds induced a G2/M block. Docking studies were carried out with a CDK1 homology model, and provide a rationale for the observed activities.     

The adjuvanticity of a mannosylated antigen reveals TLR4 functionality essential for subset specialization and functional maturation of mouse dendritic cells

The evidence that dendritic cell (DC) subsets produce differential cytokines in response to specific TLR stimulation is robust. However, the role of TLR stimulation in Ag presentation and phenotypic maturation among DC subsets is not clear. Through the adjuvanticity of a novel mannosylated Ag, mannosylated dendrimer OVA (MDO), as a pathogen-associated molecular pattern Ag, we characterized the functionality of GM-CSF/IL-4-cultured bone marrow DC and Flt3 ligand (Flt3-L) DC subsets by Ag presentation and maturation assays. It was demonstrated that both bone marrow DCs and Flt3-L DCs bound, processed, and presented MDO effectively. However, while Flt3-L CD24(high) (conventional CD8(+) equivalent) and CD11b(high) (CD8(-) equivalent) DCs were adept at MDO processing by MHC class I and II pathways, respectively, CD45RA(+) plasmacytoid DCs presented MDO poorly to T cells. Successful MDO presentation was largely dependent on competent TLR4 for Ag localization and morphological/phenotypic maturation of DC subsets, despite the indirect interaction of MDO with TLR4. Furthermore, Toll/IL-1 receptor-domain-containing adaptor-inducing IFN-beta, but not MyD88, as a TLR4 signaling modulator was indispensable for MDO-induced DC maturation and Ag presentation. Taken together, our findings suggest that DC subsets differentially respond to a pathogen-associated molecular pattern-associated Ag depending on the intrinsic programming and TLRs expressed. Optimal functionality of DC subsets in Ag presentation necessitates concomitant TLR signaling critical for efficient Ag localization and processing.     

Focus: Preventive Medicine: 2020 Evaluation of Portable Vision Screening Instruments

Amblyopia is the most common cause of preventable visual impairment in children and occurs as a result of unilateral or bilateral impairment in best-corrected visual acuity. Early diagnosis and proper treatment are crucial to prevent poor visual outcomes in adulthood. Advances in technology have provided more objective diagnostic tools, which can now be used by a wide range of healthcare providers. Here, we highlight tools that have gained popularity in the past two decades and compare clinically relevant parameters to guide primary care providers seeking to incorporate instrumental vision screening in pediatric patient care.     

Dopamine produces muscle contractions and modulates motoneuron-induced contractions inAplysia gill

1. Dopamine has been reported to exist in unusually large quantities in Aplysia gill. The physiological role of this neurotransmitter in this organ was examined. 2. The addition of dopamine to a gill perfusate results in the contractions of the lateral and medial external pinnule muscles, the circular and longitudinal muscles of the afferent vessel, and the circular muscles of the efferent vessel. 3. Dopamine-induced contractions persist after chemical synaptic transmission is eliminated in the gill. This suggests that excitatory dopamine receptors are present on gill smooth muscle fibers themselves. 4. Dopamine also potentiates the gill response to action potentials in single identified gill motoneurons. Evidence presented suggests that muscle contractions and modulation of motoneuron contractions are independent phenomena. 5. While modulation may in part be mediated by increases in excitatory junction potential (EJP) amplitude, in many cases large increases in muscle contractions occur while the enhancement of EJPs is disproportionately small. 6. Dopamine's ability to produce muscle contractions suggests that there may be dopaminergic motoneuron innervation of the gill. We suggest that dopamine's modulatory actions may be mediated via modification of excitation-contraction coupling in smooth muscle fibers.     

Inversions and other unusual heteromorphisms detected by C-banding

Summary Sixteen patients with unusual heteromorphisms involving alterations of the length and/or position of centromeric heterochromatin are described. Family studies showed that the heteromorphisms were present in other relatives and segregated in the expected 1:1 ratio. There was a significantly greater frequency of unusual heteromorphisms among Orientals than in other races studied.     

Male Horn Dimorphism and its Function in the Neotropical Dung Beetle <Emphasis Type="Italic">Sulcophanaeus velutinus</Emphasis>

Horned beetles are emerging models in the study of coevolution between novel morphologies and behavior. In Onthophagus beetles, large males use horns to fight other males in brood tunnels while small males with higher mobility sneak around the large males to gain access to females. Mating tactics have rarely been described in other dung beetle genera. We studied the horned dung beetle Sulcophanaeus velutinus that exhibits two parallel horns on the prothorax and one on the head. We put two males of different horn lengths, but similar mass, in observation chambers and found that the large male with longer horns won access to the female in physical competition. Speed tests in artificial tunnels show that locomotion is impeded in large males, suggesting an advantage in mobility for males with small horns. This work contributes to the limited existing evidence on the function of alternative morphologies in horned dung beetles taxa.     

Glucose-stimulated insulin release by individual pancreatic beta cells: potentiation by glyburide.

To investigate the effect of glyburide on insulin secretion by individual beta cells from normal rats, we employed a reverse hemolytic plaque assay. Pancreata were harvested from female Wistar-Furth rats, the pancreatic islets isolated, and the latter dispersed into single cells. These cells were mixed with protein A-coated ox erythrocytes, the mixture was placed in a Cunningham chamber in the presence of insulin antiserum, and the cells were exposed to the various test substances. Having developed hemolytic plaques around the insulin-secreting cells with complement, the percentage of plaque-forming cells was determined and the plaque areas (reflecting the amount of insulin secreted) were quantitated. For the purpose of validation, we demonstrated that (i) plaque-forming (but not nonplaque-forming) cells could be identified as insulin secreting by an independent immunofluorescent technique, (ii), plaques did not form if insulin antiserum was deleted from the preparation, (iii) plaques failed to develop if insulin antiserum was preabsorbed with insulin, and (iv) incubation with non-protein A-coated RBC or omission of complement resulted in no plaque formation. In addition, both the percentage of plaque-forming cells and the mean plaque are increased upon exposure to glucose (0.75-20 mM) in a concentration-dependent manner at 5- and 60-min incubation times. Moreover, somatostatin suppressed the percentage of plaque-forming cells and diminished the mean plaque area of cells which continued to secrete insulin in response to glucose. Exposure of cells to 100 nM glyburide in the presence of 5 mM or 20 mM glucose had no effect on the percentage of plaque-forming cells present at 5 min or 60 min. Similarly, glyburide did not alter mean plaque area at 5 or 60 min when cells were co-incubated with 5 mM glucose. However, mean plaque area was markedly enhanced at 5 and 60 min in response to glyburide and 20 mM glucose. These results demonstrate that glyburide (i) does appear to enhance insulin secretion by an effect directly on the pancreatic beta cell; (ii) does not act by recruiting previously noninsulin-secreting cells into a secretory pool; (iii) does not potentiate the effect of glucose, at fed concentrations, on insulin secretion by individual cells; but (iv) does augment insulin secretion by beta cells stimulated with supraphysiologic concentrations of glucose.