Prevalence of Human Papillomavirus Genotypes and Abnormal Pap Smears Among Women in the Military Health System.

Human papillomavirus infection (HPV) is the most common sexually transmitted infection among United States Military Servicemembers, and present in the majority of cervical cancers. Many of these infections are preventable, but HPV immunization is not mandatory during military service. The objective of this study was to examine the prevalence of vaccine-preventable cervical disease among women enrolled in the San Antonio Military Health System. This is a retrospective cross-sectional study of Pap smear results and HPV genotyping data among Military Servicewomen and beneficiaries. Simple descriptive statistics and logistic regression were used to assess the association between demographics, cervical pathology and high-risk HPV (hrHPV) infection. Pap smears were obtained by 16.9% of women and cervical pathology was present in 28.8% of samples. Compared to the 25-34 year group, 35-44 year-olds were more likely to have an abnormal Pap smear (OR 1.25, CI 1.05-1.50). Of the samples tested, 10.5% were positive for hrHPV. Adjusted multivariable analysis revealed that hrHPV infection was more likely among the 23-34 year group when compared to 35-44 (OR 0.50, CI 0.38-0.67), 45-54 (0.40. CI 0.28-0.59) and 55-65 year groups (0.46, CI 0.30-0.71). Active Duty Servicewomen were more likely to test positive for hrHPV when compared to Active Duty Family Members (OR 0.59, CI 0.45-0.79) and Retiree Family Members (OR 0.59, CI 0.41-0.83). Younger women and Active Duty Servicewomen are significantly more likely to have cervical infection with hrHPV. Future studies should assess the cost-effectiveness of mandatory HPV immunization for military members.

Oxygen reduction reaction induced pH-responsive chemo-mechanical hydrogel actuators.

We describe and characterize elementary designs for electrochemical micro- and macro-scale chemomechanical hydrogel actuators. The actuation of a pH-sensitive cross-linked polyacrylic acid (PAA) hydrogel is driven in the model devices through the oxygen reduction reaction (ORR) occurring at the electrodes of an embedded Au mesh micro-electrochemical array. Proton consumption by the ORR at the cathode of the embedded electrochemical cell leads to the formation of a localized pH gradient that in turn drives the strain response in the composite actuators. The dynamics result from the ionization of the carboxylic acid moieties of the PAA network in the high pH region, yielding an osmotic pressure that drives a volumetric expansion due to water imbibition. This system actuates both stably and reversibly; when the electrochemically-induced ORR is halted, the localized pH gradient dissipates due to diffusive mixing, which in turn relaxes the induced strains. Two approaches to the fabrication of hydrogel actuators were examined in this work. The first method adopted a design based on small flagella (∼0.2 mm × 1.5 mm × 60 µm, width × length × height) in which the actuating PAA structures are molded atop a set of fixed electrodes mounted on a supporting substrate. These hydrogel actuators show fast, large-amplitude, and largely reversible responses in the ORR mediated chemomechanical dynamics. We also investigated larger hydrogel actuators (∼4.5 mm × 11 mm × 1 mm, width × length × height), based on an autonomous design that embeds an open mesh stretchable micro-electrode array within the hydrogel. The significant and design-dependent impacts of mass transfer on the chemomechanical dynamics are evidenced in each case, a feature examined to elucidate more efficient mesoscopic design rules for actuators of this form.

Chromatin configuration and epigenetic landscape at the sex chromosome bivalent during equine spermatogenesis.

Pairing of the sex chromosomes during mammalian meiosis is characterized by the formation of a unique heterochromatin structure at the XY body. The mechanisms underlying the formation of this nuclear domain are reportedly highly conserved from marsupials to mammals. In this study, we demonstrate that in contrast to all eutherian species studied to date, partial synapsis of the heterologous sex chromosomes during pachytene stage in the horse is not associated with the formation of a typical macrochromatin domain at the XY body. While phosphorylated histone H2AX (γH2AX) and macroH2A1.2 are present as a diffuse signal over the entire macrochromatin domain in mouse pachytene spermatocytes, γH2AX, macroH2A1.2, and the cohesin subunit SMC3 are preferentially enriched at meiotic sex chromosome cores in equine spermatocytes. Moreover, although several histone modifications associated with this nuclear domain in the mouse such as H3K4me2 and ubH2A are conspicuously absent in the equine XY body, prominent RNA polymerase II foci persist at the sex chromosomes. Thus, the localization of key marker proteins and histone modifications associated with the XY body in the horse differs significantly from all other mammalian systems described. These results demonstrate that the epigenetic landscape and heterochromatinization of the equine XY body might be regulated by alternative mechanisms and that some features of XY body formation may be evolutionary divergent in the domestic horse. We propose equine spermatogenesis as a unique model system for the study of the regulatory networks leading to the epigenetic control of gene expression during XY body formation.

Cloning and characterization of novel human SLC4A8 gene products encoding Na+-driven Cl-/HCO3(-) exchanger variants NDCBE-A, -C, and -D.

The reported sequences of the human and mouse Na+-driven Cl-/HCO3(-) exchangers (NDCBEs) differ greatly in their extreme cytosolic COOH termini (Ct). In human NDCBE (NDCBE-B), a 17-amino acid (aa) sequence replaces 66 aa at the equivalent position in mouse NDCBE (NDCBE-A). We performed 5'- and 3'-rapid amplification of cDNA ends (RACE) on human brain cDNA, followed by PCR of full-length cDNAs to determine whether the human SLC4A8 gene was capable of producing the mouselike Ct sequence. Our study confirmed the presence in human cDNA of mouse NDCBE-like transcripts (human NDCBE-A) and also disclosed the existence of three further novel NDCBE transcripts that we have called NDCBE-C, NDCBE-D, and NDCBE-D'. The novel NDCBE-C/D/D' transcripts initiate at a novel "exon 0" positioned approximately 35 kb upstream of the first exon of NDCBE-A/B. NDCBE-C/D/D' protein products are predicted to be truncated by 54 aa in the cytosolic NH(2) terminus (Nt) compared with NDCBE-A/B. Our data, combined with a new in silico analysis of partial transcripts reported by others in the region of the human SLC4A8 gene, increase the known extent of the SLC4A8 gene by 49 kb, to 124 kb. A functional comparison of NDCBE-A/B/C/D expressed in Xenopus oocytes demonstrates that the Nt variation does not affect the basal functional expression of NDCBE, but those with the shorter Ct have a 25-50% reduced functional expression compared with those with the longer Ct. By comparison with an artificially truncated NDCBE that contains neither 17-aa nor 66-aa Ct cassette, we determined that the functional difference is unrelated to the 66-aa cassette of NDCBE-A/C, but is instead due to an inhibitory effect of the 17-aa cassette of NDCBE-B/D.

Characterization of human SLC4A10 as an electroneutral Na/HCO3 cotransporter (NBCn2) with Cl- self-exchange activity.

The SLC4A10 gene product, commonly known as NCBE, is highly expressed in rodent brain and has been characterized by others as a Na(+)-driven Cl-HCO(3) exchanger. However, some of the earlier data are not consistent with Na(+)-driven Cl-HCO(3) exchange activity. In the present study, northern blot analysis showed that, also in humans, NCBE transcripts are predominantly expressed in brain. In some human NCBE transcripts, splice cassettes A and/or B, originally reported in rats and mice, are spliced out. In brain cDNA, we found evidence of a unique partial splice of cassette B that is predicted to produce an NCBE protein with a novel C terminus containing a protein kinase C phosphorylation site. We used pH-sensitive microelectrodes to study the molecular physiology of human NCBE expressed in Xenopus oocytes. In agreement with others we found that NCBE mediates the 4,4'-diisothiocyanato-stilbene-2,2'-disulfonic acid-sensitive, Na(+)-dependent transport of HCO(3)(-). For the first time, we demonstrated that this transport process is electroneutral. Using Cl(-)-sensitive microelectrodes positioned at the oocyte surface, we found that, unlike both human and squid Na(+)-driven Cl-HCO(3) exchangers, human NCBE does not normally couple the net influx of HCO(3)(-) to a net efflux of Cl(-). Moreover we found that that the (36)Cl efflux from NCBE-expressing oocytes, interpreted by others to be coupled to the influx of Na(+) and HCO(3)(-), actually represents a CO(2)/HCO(3)(-)-stimulated Cl(-) self-exchange not coupled to either Na(+) or net HCO(3)(-) transport. We propose to rename NCBE as the second electroneutral Na/HCO(3) cotransporter, NBCn2.

A conductive pathway generated from fragments of the human red cell anion exchanger AE1.

Human red cell anion exchanger AE1 (band 3) is an electroneutral Cl-HCO3- exchanger with 12-14 transmembrane spans (TMs). Previous work using Xenopus oocytes has shown that two co-expressed fragments of AE1 lacking TMs 6 and 7 are capable of forming a stilbene disulphonate-sensitive (36)Cl-influx pathway, reminiscent of intact AE1. In the present study, we create a single construct, AE1Delta(6: 7), representing the intact protein lacking TMs 6 and 7. We expressed this construct in Xenopus oocytes and evaluated it employing a combination of two-electrode voltage clamp and pH-sensitive microelectrodes. We found that, whereas AE1Delta(6: 7) has some electroneutral Cl-base exchange activity, the protein also forms a novel anion-conductive pathway that is blocked by DIDS. The mutation Lys(539)Ala at the covalent DIDS-reaction site of AE1 reduced the DIDS sensitivity, demonstrating that (1) the conductive pathway is intrinsic to AE1Delta(6: 7) and (2) the conductive pathway has some commonality with the electroneutral anion-exchange pathway. The conductance has an anion-permeability sequence: NO3- approximately I- > NO2- > Br- > Cl- > SO4(2-) approximately HCO3- approximately gluconate- approximately aspartate- approximately cyclamate-. It may also have a limited permeability to Na+ and the zwitterion taurine. Although this conductive pathway is not a usual feature of intact mammalian AE1, it shares many properties with the anion-conductive pathways intrinsic to two other Cl-HCO3- exchangers, trout AE1 and mammalian SLC26A7.

Effect of human carbonic anhydrase II on the activity of the human electrogenic Na/HCO3 cotransporter NBCe1-A in Xenopus oocytes.

Others report that carbonic anhydrase II (CA II) binds to the C termini of the anion exchanger AE1 and the electrogenic Na/HCO3 cotransporter NBCe1-A, enhancing transport. After injecting oocytes with NBCe1-A cRNA (Day 0), we measured NBC current (I(NBC)) by two-electrode voltage clamp (Day 3), injected CA II protein + Tris or just Tris (Day 3), measured I(NBC) or the initial rate at which the intracellular pH fell (dpH(i)/dt) upon applying 5% CO2 (Day 4), exposed oocytes to the permeant CA inhibitor ethoxzolamide (EZA), and measured I(NBC) or dpH(i)/dt (Day 4). Because dpH(i)/dt was greater in CA II than Tris oocytes, and EZA eliminated the difference, injected CA II was functional. I(NBC) slope conductance was unaffected by injecting CA II. Moreover, EZA had identical effects in CA II versus Tris oocytes. Thus, injected CA II does not enhance NBC activity. In a second protocol, we made a fusion protein with enhanced green fluorescent protein (EGFP) at the 5' end of NBCe1-A and CA II at the 3' end (EGFP-e1-CAII). We measured I(NBC) or dpH(i)/dt (days 3-4), exposed oocytes to EZA, and measured I(NBC) or dpH(i)/dt (Day 3-4). dpH(i)/dt was greater in oocytes expressing EGFP-e1-CA II versus EGFP-e1, and EZA eliminated the difference. Thus, fused CA II was functional. Slope conductances of EGFP-e1-CAII versus EGFP-e1 oocytes were indistinguishable, and EZA had no effect. Thus, even when fused to NBCe1-A, CA II does not enhance NBCe1-A activity.

The human NBCe1-A mutant R881C, associated with proximal renal tubular acidosis, retains function but is mistargeted in polarized renal epithelia.

The human electrogenic renal Na-HCO(3) cotransporter (NBCe1-A; SLC4A4) is localized to the basolateral membrane of proximal tubule cells. Mutations in the SLC4A4 gene cause an autosomal recessive proximal renal tubular acidosis (pRTA), a disease characterized by impaired ability of the proximal tubule to reabsorb HCO(3)(-) from the glomerular filtrate. Other symptoms can include mental retardation and ocular abnormalities. Recently, a novel homozygous missense mutant (R881C) of NBCe1-A was reported from a patient with a severe pRTA phenotype. The mutant protein was described as having a lower than normal activity when expressed in Xenopus oocytes, despite having normal Na(+) affinity. However, without trafficking data, it is impossible to determine the molecular basis for the phenotype. In the present study, we expressed wild-type NBCe1-A (WT) and mutant NBCe1-A (R881C), tagged at the COOH terminus with enhanced green fluorescent protein (EGFP). This approach permitted semiquantification of surface expression in individual Xenopus oocytes before assay by two-electrode voltage clamp or measurements of intracellular pH. These data show that the mutation reduces the surface expression rather than the activity of the individual protein molecules. Confocal microscopy on polarized mammalian epithelial kidney cells [Madin-Darby canine kidney (MDCK)I] expressing nontagged WT or R881C demonstrates that WT is expressed at the basolateral membrane of these cells, whereas R881C is retained in the endoplasmic reticulum. In summary, the pathophysiology of pRTA caused by the R881C mutation is likely due to a deficit of NBCe1-A at the proximal tubule basolateral membrane, rather than a defect in the transport activity of individual molecules.