Frequent detection of HIV-1 RNA but low rates of HIV-1 isolation in cervicovaginal secretions from infected women

Information regarding the presence of HIV-1 in the female genital tract is necessary to gain insight into the mechanism of HIV-1 heterosexual transmission. Herein, we present the results of a study on virus isolation and HIV-1 RNA detection from cervicovaginal lavage (CVL) samples from 25 HIV-1 seropositive women. Despite detectable levels of HIV-1 RNA in 88% of CVL samples, HIV-1 was isolated in only four (19%) samples. Although HIV-1 shedding in cervicovaginal secretions is a common event at all disease stages, the recovery of infectious virus in cell cultures appears to be rare; this renders viral isolation in studies aimed to evaluate the infectivity of cervicovaginal secretions relatively useless.     

Limited secretory-IgA response in cervicovaginal secretions from HIV-1 infected, but not high risk seronegative women: lack of correlation to genital viral shedding

The presence and antigen specificity of IgG and secretory-IgA (s-IgA) to HIV-1 were evaluated in cervicovaginal lavages (CVL) from 26 infected and 10 high-risk seronegative women. All the seropositive women had detectable IgG recognizing several viral antigens, while a smaller percentage of women demonstrated s-IgA to the virus. In addition, s-IgA were of limited specificity and provided weak reactivities on Immunoblot bands; an almost constant absence of s-IgA to gp120 was also observed. Neither the presence nor the specificity of either IgG or s-IgA to the virus in CVL prevented the shedding of HIV-1 in this body fluid; in fact, viral RNA was detected in all the women studied and the amounts of viral shedding was unrelated to the genital antibody response. On the other hand, none of the high-risk seronegative women had detectable antibodies to HIV-1 in CVL of either the IgG or s-IgA isotype. Our results a) confirm an impairment of mucosal antibody response during HIV-1 infection and suggest that mucosal immunity is not able to prevent viral shedding in the female genital tract and thus cannot modulate the infectivity of genital secretions; aa) do not provide evidence for a mucosal "memory/protective" antibody response in the genital tract of high-risk seronegative women.     

Nearest-neighbor relationships of the constituent polypeptides in plastoquinol-plastocyanin oxidoreductase

The nearest-neighbor relationship among the constituent polypeptides of the isolated plastoquinol-plastocyanin oxidoreductase from spinach chloroplasts has been investigated. (1) The isolated plastoquinol-plastocyanin oxidoreductase (the b6/f complex) is treated with various concentrations of the cross-linker glutaraldehyde. The treated b6/f complexes are then analyzed by SDS-polyacrylamide gel electrophoresis coupled with the immunodecoration of cross-link products by specific antibodies for each of the four prominent constituent polypeptides. Cytochrome b6 is found to be most resistant to forming any intermolecular cross-link products. At low concentrations of glutaraldehyde, the 'Rieske' iron-sulfur (Fe-S) protein and subunit IV of the b6/f complex, however, appear to form cross-link products with a relative molecular weight of 35 000. Dimers of cytochrome f and cytochrome f/Rieske protein cross-link products can also be detected. (2) When a Rieske Fe-S protein-depleted b6/f complex is used in place of the control b6/f complex, cytochrome b6 is less resistant to intermolecular cross-linking, while subunit IV does not form any 35 kDa cross-link product, unlike the case in control b6/f complex. Subunit IV is concluded to be closely associated with the Rieske Fe-S protein. This provides evidence that subunit IV is a bona fide component of the cytochrome b6/f complex, although no function can yet be assigned to it. The results are discussed in relationship to the spatial and functional relationships among the components of the b6/f complex.     

The optical activity of polypeptides

The present status of the theory of the optical properties of polypeptides is discussed in a general way. Arguments are presented that indicate that the interaction of exciton bands in helices will occur in a tensorial way with the parallel polarized bands normally having stronger interactions than the perpendicular polarized bands, with only small cross-interactions. The net result is an anisotropy of the hyperchromism of the ππ* band and an imbalance of the rotatory strengths of the parallel and perpendicular components. According to the evidence of Mandel and Holzwarth [(1973) Biopolymers 13 , 655–674], this is, in fact, the origin of the strongly nonconservative CD bands of polyproline II and related structures. The implications of anisotropic hyperchromicity on the interpretation of the linear dichroism of crystals is discussed.     

Estradiol and progesterone regulation of immunoglobulin A and G and secretory component in cervicovaginal secretions of the rat

The present study examined the influence of hormones on the levels of immunoglobulins A (IgA) and G (IgG) and secretory component (SC) in cervicovaginal secretions of ovariectomized rats. Administration of estradiol to ovariectomized rats resulted in a significant decline in cervicovaginal content of IgA, IgG and SC. This response was dose dependent and was not prevented by administration of dexamethasone, a potent synthetic glucocorticoid, with estradiol. Treatment of ovariectomized rats with progesterone also lowered the levels of IgA and SC in cervicovaginal secretions. In contrast, dexamethasone had no apparent vaginal effect. The action of estradiol on cervicovaginal IgA, IgG and SC appears to be independent of uterine influence. This conclusion is based upon our observation that estrogen treatment of rats with ligations at their uterocervical junction still have decreased cervicovaginal IgA and SC levels. In parallel with this inhibitory effect, estradiol administration stimulated the accumulation of IgA and SC in uterine secretions. These findings indicate that the sex hormones play a role in regulating IgA, IgG and SC content in cervicovaginal secretions. In addition, it suggests that hormonal balance in females may influence the immune response of the reproductive tract to infectious disease.     

The induction of large unilamellar vesicle fusion by cationic polypeptides: the effects of mannitol,size, charge density and hydrophobicity of the cationic polypeptides

Polylysine and lysine-based copolymers induced fusion of large unilamellar vesicles only in media containing at least 0.4 M mannitol. In the absence of mannitol, polylysine and certain lysine-based copolymers also containing acidic amino acids were not able to induce fusion. Fusion, in the presence of mannitol, was induced at nanomolar concentrations of the polycations. Excess polymer caused reduced rate and extent of fusion. In the presence of 100 mM NaCl the effective concentration range of the polycations was narrower. Kinetic analysis determined that salt increased the aggregation constant C₁₁ while reducing the fusion constant f₁₁. Addition of polylysine in excess resulted in smaller C₁₁. Short polylysine (3500) was less effective on a molar basis than a long one (37 000). Copolymers were more effective than polylysine due to higher aggregation potential. Copolymers were also more effective in promoting Ca²⁺-induced fusion in the absence of mannitol, their greater efficiency resulting from substantially larger fusion potential, without a greater rate of leakage. Preincubation of the vesicles with the polycations for less than 20 s resulted in faster fusion rates, while longer preincubations caused slower fusion rates. Addition of polycations to the preincubated mixture enhanced the fusion rates, indicating that the polycations were not available, rather than the vesicles being not susceptible to fusion. The effect of preincubation suggests two phases in the binding of the polycations to the vesicles; a fast phase of partial binding and a slower phase resulting in complete binding. The addition of millimolar concentrations of pyrophosphate or sulphate provided a fine control of the effective polycation concentration and its effect on fusion.     

Improved non-chromatographic purification of a recombinant protein by cationic elastin-like polypeptides

This paper reports an improvement in the purification of thioredoxin (Trx) expressed from E. coli by inverse transition cycling (ITC) using cationic elastin-like polypeptides (ELPs). Two ELP libraries having 2% and 5% lysine residues and molecular weights ranging from 4 to 61.1 kDa showed greater salt sensitivity in their inverse transition behavior than purely aliphatic ELPs. Expression yield of Trx-ELP fusions was an unpredictable function of guest residue composition, but reducing the molecular weight of the ELP tag generally increased Trx yield. A cationic 4.3 kDa ELP is the shortest ELP used to purify any protein by ITC to date. A 15.9 kDa ELP with a guest residue composition of K:V:F of 1:7:1 was found to be the optimal cationic tag to purify Trx, as it provided 50% greater Trx yield and only required one-fifth the added NaCl for purification of Trx as compared to previously used aliphatic ELP tags.     

Superoxide dismutase activity of digestive secretions

A superoxide dismutase (SOD) activity in saliva and gastric juice has been revealed using the NBT reduction and adrenaline autooxidation assays. This activity in saliva is induced mainly by enzymic mechanisms, whereas in gastric juice--by nonenzymic mechanisms. The gel chromatographic analysis of saliva has demonstrated the presence of several SOD isoenzymes. Nonenzymic mechanisms of the SOD activity in gastric juice involve the ionic activity of H+, Cu2+ and that of reduced glutathione. The SOD activity in gastric juice can be decreased by adding antacids and Fe ions. The effect of Fe3+ is found to be different depending on the methods of the SOD activity evaluation. Such metal-chelating agents as histidine and organic acids increase the SOD activity in gastric juice.     

Antiprotease activity in tears and nasal secretions

Summary A trypsin-inhibitory capacity in tears and nasal secretions is demonstrated. No correlation exists between the serum a₁-AT genotype and the level of trypsin-inhibitory capacity in these secretions. Application of the radial-immunodiffusion technique indicates that the antiprotease activity in tears is different from that associated with a₁ globulin.

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Zusammenfassung Im Nasen- und Tränensekret gelang der Nachweis von Antiproteasen-aktivität (Trypsin-Hemmkapazität). Zwischen dem Serum a₁-AT-Genotyp und dem Ausmaß der Trypsin-Hemmkapazität in diesen Sekreten besteht keine Korrelation. Mittels der Radial-Immunodiffusionstechnik konnte gezeigt werden, daß die Antiproteasenkapazität im Tränensekret immunologisch nicht mit dem Serum-a₁-Antitrypsin identisch ist.

     

1964例围产期孕妇阴道分泌物常见病原体分析

目的探讨围产期孕妇阴道分泌物常见病原体的感染状况,减少不良妊娠的发生。方法假丝酵母菌(C)、滴虫(T)检测采用涂片法,细菌性阴道炎(BV)检测采用唾液酸酶法,淋球菌(NG)检测采用平板培养法,解脲支原体(UU)检测采用人工培养法,沙眼衣原体(CT)检测采用乳胶免疫层析法。结果 1 964例围产期孕妇中病原体感染者共360例,占总人数的18.33%(C 9.41%、T 0.46%、BV 6.36%、NG 0.00%、UU 4.53%、CT 1.73%)。其中单一病原体感染者为281例,占总人数的14.31%;两种或两种以上病原体感染者为79人,占总人数的4.02%。结论抚顺地区围产期孕妇阴道分泌物病原体构成复杂,病原体主要以假丝酵母菌和细菌性阴道炎为主,其次是解脲支原体,部分孕妇可同时感染多种病原体。     

妊娠妇女与无性生活女性阴道分泌物微生物组学分析

目的 探讨妊娠妇女与无性生活女性阴道分泌物中菌群结构差异,为临床女性泌尿生殖系统疾病研究提供可靠的依据。方法 运用高通量二代测序技术检测20例妊娠妇女和29例无性生活女性阴道分泌物中微生物;采用秩和检验进行组间显著性差异分析,使用生物信息学软件进行数据处理。结果 49份样本共注释出14个门、23个纲、33个目、28个科、38个属和23个种水平物种。妊娠妇女与无性生活女性阴道分泌物中菌群结构差异显著。妊娠妇女在种水平仅有Atopobium vaginae、Campylobacter ureolyticus、Lactobacillus coleohominis、德氏乳杆菌、瑞氏乳杆菌、惰性乳杆菌、罗伊乳杆菌、白假丝酵母和近平滑假丝酵母共9个物种。结论 妊娠能引起女性阴道分泌物菌群结构发生改变,且菌种数量明显减少。Lactobacillus coleohominis和德氏乳杆菌可能在维持妊娠妇女阴道菌群平衡和自净作用中起到关键性作用。     

Immunological cross-reactivity of multiplication-stimulating activity polypeptides

The immunoreactivity of the multiple species of multiplication-stimulating activity (MSA) purified from medium conditioned by a rat liver cell line (BRL-3A) has been examined. Antibodies were raised in rabbits following immunization with MSA II polypeptides. Subpopulations of antibodies were purified from one antiserum using DEAE-cellulose chromatography. One antibody subpopulation recognized common antigenic determinants on MSA I and MSA II polypeptides; whereas a second antibody subpopulation recognized common determinants on MSA I, II, and III polypeptides. In a radioimmunoassay utilizing 125I-MSA III-2 and a purified antibody subpopulation, the human somatomedins (somatomedin A, insulin-like growth factor I and II) showed weak, but significant cross-reactivity: insulin-like growth factor II was 10% as potent as MSA II. By contrast, somatomedin partially purified from rat serum, insulin, growth hormone, epidermal growth factor, nerve factor, and fibroblast growth factor, showed no reactivity in the radioimmunoassay.     

Optical activity of polypeptides and proteins

Using methods described in a previous publication the optical activity of a number of polypeptides and proteins has been calculated. The systems included the α-helix, the two β-structures, polyproline I, polyproline II, collagen and collagen models, and poly-N-methylalanine. In addition to these orderded structures, calculations were also performed on the α, β and nonperiodic regions of myoglobin, lysozyme, ribonuclease-S and β-chymotrypsin. The α and β structures in prteins differ from the polypeptide models by being very short and partially disordered. It is concluded that the 222-nm band of the α-helix is a good method for detecting helices in proteins but that the 207-and 191-nm bands of the helix will not fit a linear superposition model. The circular dichroism of the so-called β regions of proteins differs markedly from that for ideal β structure because of breakdownin symmetry. As a result estimates of β-structure in proteins based on polypeptide models are not likely to be quantitative. The theoretical methods give an adequate account of the optical activity of all the ordered polypeptides except polyproline II and collagen and (by inference) the nonperiodic chains in the various proteins. This difficulty is the remaining barrier to a complete theory of the optical activity of the polypeptide backbone in globular proteins.     

Enhancement of UDPgalactose: glycoprotein galactosyltransferase in cultured human skin fibroblasts by cationic polypeptides

UDPgalactose : glycoprotein galactosyltransferase in normal human skin fibroblast homogenates has been assayed using ovalbumin as an acceptor. The activity in the homogenate fraction sedimenting between 51 300 X g and 105 000 X g was enhanced by the addition of a number of catonic polypeptides of L-configuration but not by those of D-configuration. In contrast to the enhancing effect of poly(L-lysine), poly(L-glutamic acid) inhibited the activity. Poly(D-glutamic acid) had no effect. Cationic or anionic amino acid derivatives, spermine or spermidine had no effect on activity. The enhancement of transferase activity by poly(L-arginine) is probably due to an increase in V for UDPgalactose and ovalbumin. The implication of these results for the regulation of glycoprotein synthesis in cultivated skin fibroblasts and for the pathogenesis of cystic fibrosis is discussed.     

Inhibition of immune aggregate-induced activation of the first complement component by cationic polypeptides

A cationic amino acid copolymer (CP530) with a molar ratio of lysine, leucine, tryptophan and phenylalanine of 11:2:1:1 and a M_r of about 2300 was prepared and its inhibitory effects on the complement cascade was compared with those of polylysine with a M_r of about 3000. The effects of these two cationic peptides appeared to be at the early stage of complement activation. CP530 and polylysine inhibited the binding of C1q to insoluble IgG aggregates with a concentration required for 50% inhibition of 0.7 and 0.9 mM, respectively. Both compounds were also potent inhibitors of immune hemolysis (a concentration causing 50% inhibition, 0.5 and 3.5 μM respectively) as well as well as assembly of EAC cell intermediates required for formation of C3 and C5 convertases (a concentration for 50% inhibition of 1.0 μM for CP530 and 3.8 μM for polylysine). However, CP530 was shown to be distinctly more effective against the activation of C1r·Cls complex induced by insoluble IgG aggregate-bound C1q, requiring 0.15 mM for 50% inhibition compared to greater than 10 mM for polylysine. The 50% inhibition value for soluble IgG aggregate-induced activation of C1 in whole serum was 0.7 mM for CP530 and 5.0 mM for polylysine. The greater the inhibition of C1 activation by CP530 than that exerted by polylysine could be attributable to the presence of non-lysyl residues which provide the structural basis for specificity and potency.