Immunolocalization of inhibin α-subunit in the human testis

Summary The localization of inhibin -subunit in the human testis was studied at the light- and electron-microscope level with immunostaining techniques. Antibodies against specific fragments of porcine and human inhibin -subunits were utilized. At light microscopy, inhibin -subunit immunoreactivity was detected in Sertoli cells, spermatocytes and in some Leydig cells. At electron microscopy, gold labeling was found in the cisternae of the Golgi apparatus and in the endoplasmic reticulum of Sertoli and Leydig cells. Gold labeling for inhibin was also found in coated vesicles in the cytoplasm of Sertoli cells as well as in coated pits and coated vesicles in the cytoplasm of some spermatocytes. The results of the present study suggest that, in the human testis, inhibin is produced by Sertoli and Leydig cells and is taken up by spermatocytes, on which it might act in a paracrine manner.     

Morphology of the “spindle-shaped body” in the developing tail of human spermatids

Summary At the beginning of the late cap phase of the normal differentiation of human spermatids, a unique structure, the spindle shaped body, occupies part of the future middlepiece. This spindle-shaped body was followed throughout its development, and its functional relationship to the formation of the fibrous sheath is discussed. The spindle-shaped body disappears when the fibrous sheath is completed and the annulus moves down, giving rise to the middle-piece.Supported by Grants from the Deutsche Forschungsgemeinschaft.     

Role of A-chain in functioning of the active site of human α-thrombin

This review summarizes current data suggesting that A-chain of the human alpha-thrombin molecule plays a role of allosteric effector in catalytic reactions with various substrates. Special attention is paid to the relationship between A-chain structure and catalytic activity of thrombin. The existence of this relationship is based on studies of natural mutation of A-chain of the alpha-thrombin molecule. Use of molecular and essential dynamics confirmed the role of A-chain in changes of conformation and catalytic properties of this enzyme; these changes involve residues located in the specificity sites and some inserting loops. Current knowledge on structure and properties of thrombin can be used for the development of new antithrombin agents.     

In silico prediction of human pathogenicity in the γ-proteobacteria

Background

Although the majority of bacteria are innocuous or even beneficial for their host, others are highly infectious pathogens that can cause widespread and deadly diseases. When investigating the relationships between bacteria and other living organisms, it is therefore essential to be able to separate pathogenic organisms from non-pathogenic ones. Using traditional experimental methods for this purpose can be very costly and time-consuming, and also uncertain since animal models are not always good predictors for pathogenicity in humans. Bioinformatics-based methods are therefore strongly needed to mine the fast growing number of genome sequences and assess in a rapid and reliable way the pathogenicity of novel bacteria.

Methodology/Principal Findings

We describe a new in silico method for the prediction of bacterial pathogenicity, based on the identification in microbial genomes of features that appear to correlate with virulence. The method does not rely on identifying genes known to be involved in pathogenicity (for instance virulence factors), but rather it inherently builds families of proteins that, irrespective of their function, are consistently present in only one of the two kinds of organisms, pathogens or non-pathogens. Whether a new bacterium carries proteins contained in these families determines its prediction as pathogenic or non-pathogenic. The application of the method on a set of known genomes correctly classified the virulence potential of 86% of the organisms tested. An additional validation on an independent test-set assigned correctly 22 out of 24 bacteria.

Conclusions

The proposed approach was demonstrated to go beyond the species bias imposed by evolutionary relatedness, and performs better than predictors based solely on taxonomy or sequence similarity. A set of protein families that differentiate pathogenic and non-pathogenic strains were identified, including families of yet uncharacterized proteins that are suggested to be involved in bacterial pathogenicity.     

Contribution of gene conversion in the evolution of the human β-like globin gene family

Gene conversion is referred to as one of two types of mechanisms known to act on gene families, mainly to maintain their sequence homogeneity or, in certain cases, to produce sequence diversity. The concept of gene conversion was established 20 years ago by researchers working with fungi. A few years later, gene conversion was also observed in the human genome, i.e. the γ-globin locus. The aim of this article is to emphasize the role of genetic recombination, particularly of gene conversion, in the evolution of the human β-like globin genes and further to summarize its contribution to the convergent evolution of the fetal globin genes. Finally, this article attempts to re-examine the origin and spread of specific mutations of the β-globin cluster, such as the sickle cell or β-thalassemia mutations, on the basis of repeated gene conversion events. Received: 13 February 1997 / Accepted: 15 May 1998     

Studies on the metabolism of steroids in the foetus. Biosynthesis of 6α-hydroxytestosterone in the human foetal liver

After incubation of testosterone with 105000g microsomes of human foetal liver, 6alpha-hydroxytestosterone was isolated and identified by t.l.c. and g.l.c.-mass spectrometry. This is the first example of 6alpha-hydroxylation of C(19) steroids in the human liver, and the finding is discussed in relation to earlier reports of 6-oxygenated C(19) and C(18) steroids in pregnant women.     

Pitfalls in the quantitative estimation of β-amyloid immunoreactivity in human brain tissue

 The duration of formic acid (FA) pretreatment clearly influences the extent of β-amyloid immunoreactivity in brain tissue and consequently also the results of quantitative analysis. All of the parameters studied (area fraction, density, and mean size of β-amyloid deposits) significantly increased with pretreatment of up to 6 h with β-amyloid antibody obtained from Dako. Longer exposure to FA only marginally increased the mean size of the single deposits, whereas the area fraction and the density of β-amyloid deposits slightly decreased. Optimal 6-h pretreatment (or even longer) did not reveal any β-amyloid aggregates in those cases where none was seen with shorter durations of FA pretreatment. Similar results were obtained with β-amyloid antibody 4G8 obtained from Senetek, whereas β-amyloid antibody 6E10 was shown to be less dependent upon FA pretreatment. In conclusion, we recommend that the FA pretreatment time should be studied and optimized for each antibody used and always be described when the quantitative analysis of β-amyloid load is reported. Accepted: 7 April 1998     

Assessing spatiotemporal associations in the occurrence of badger–human conflict in England

Examples from a variety of taxa demonstrate that under certain circumstances, the exclusion or translocation of ‘problem’ animals is ineffective in resolving human–wildlife conflicts and may even elicit new problems elsewhere. Damage caused by badger setts (burrows) is an important source of human–wildlife conflict in the UK and is commonly managed by excluding badgers from all or part of problem setts. We used records of licences issued for the management of such problems and a novel statistical approach to assess spatiotemporal associations between problem cases in England from 2002 to 2005. We predicted that management at urban badgers' setts, and particularly exclusion of badgers from urban main setts, would give rise to subsequent problems at focal setts and in neighbouring areas. Frequencies of problems occurring at individual setts were similar in urban and rural areas. In areas neighbouring setts subjected to management action, the background frequency of problems was higher in urban than in rural areas, reflecting the occurrence of problems at a higher proportion of urban setts. The frequency of new cases arising at or in the vicinity of managed setts within a critical time period after management action was not significantly different from the background frequency of problems for any combination of land use, sett type and management approach. This finding suggests that the measures currently employed for managing problem setts do not importantly increase the likelihood of problems reoccurring in the same location or emerging nearby.     

An expression profile of human α-lactalbumin in the milk of transgenic mouse

Five female transgenic mice were produced by microinjection using a construct made up of a 7.3-kb-5′ flanking region and a 2.0-kb coding region of human α-lactalbumin, as well as a 227-bp 3′-flanking region from bovine growth hormone gene. A founder female expressed human α-lactalbumin as much as 0.3 g per liter of its milk, approximately a 3-fold increase in the total α-lactalbumin concentration of the transgenic mouse milk. Compared with the normal mice, the expression profile of thehα-Lac transgene in the transgenics is different during the lactation, showing low level in the first 3 days and becoming increased from day 4, then gradually reaching and stabilizing at the highest level from day 13. In addition, the milk yielding volume in the transgenics tended to be higher than in normal mice, suggesting higher concentrations of α-lactalbumin might boost more milk output.     

An expression profile of human α-lactalbumin in the milk of transgenic mouse

Five female transgenic mice were produced by microinjection using a construct made up of a 7.3-kb-5′ flanking region and a 2.0-kb coding region of human α-lactalbumin, as well as a 227-bp 3′-flanking region from bovine growth hormone gene. A founder female expressed human α-lactalbumin as much as 0.3 g per liter of its milk, approximately a 3-fold increase in the total α-lactalbumin concentration of the transgenic mouse milk. Compared with the normal mice, the expression profile of the hα-Lac transgene in the transgenics is different during the lactation, showing low level in the first 3 days and becoming increased from day 4, then gradually reaching and stabilizing at the highest level from day 13. In addition, the milk yielding volume in the transgenics tended to be higher than in normal mice, suggesting higher concentrations of α-lactalbumin might boost more milk output.     

An expression profile of human α-lactalbumin in the milk of transgenic mouse

a-lactalbumin(a-Lac),amajorwheyprotein,isacalciummetalloprotein,thathasbeenfoundinallmilksstudiedsofar.ItinteractswithUDP-galactosyl-transferasetoformthelactosesynthetaseandthusmightbeakeyproteinforlactogenesis.Lactosesyn-thetaseispostulatedtobetherate-limitingenzymeforlactosebiosynthesis.Theincreaseda-Lacactivitycanproducesufficientlactosesynthetaseforthesynthesisoflactose,andinmilkyieldbydrawingwaterintomilk,sincelactoseisanosmoreactivemolecule.Transgenicswineoverexpressingbovinea-lactalbu…     

Detection of pro-apoptotic Bax∆2 proteins in the human cerebellum

Bax?2 is a pro-apoptotic protein originally discovered in colon cancer patients with high microsatellite instability. Unlike most pro-apoptotic Bax family members, Bax?2 mediates cell death through a non-mitochondrial caspase 8-dependent pathway. In the scope of analyzing the distribution of Bax?2 expression in human tissues, we examined a panel of human brain samples. Here, we report four cerebellar cases in which the subjects had no neurological disorder or disease documented. We found Bax?2 positive cells scattered in all areas of the cerebellum, but most strikingly concentrated in Purkinje cell bodies and dendrites. Two out the four subjects tested had strong Bax?2-positive staining in nearly all Purkinje cells; one was mainly negative; and one had various levels of positive staining within the same sample. Further genetic analysis of the Purkinje cell layer, collected by microdissection from two subjects, showed that the samples contained G7 and G9 Bax microsatellite mutations. Both subjects were young and had no diseases reported at the time of death. As the distribution of Bax?2 is consistent with that known for Baxα, but in a less ubiquitous manner, these results may imply a potential function of Bax?2 in Purkinje cells.     

Secretory expression of the α-subunit of human coagulation factor XIII in the yeast Pichia pastoris

Pro-FXIIIa (the -subunit of FXIII with activation peptide, which must be removed to produce the active form of FXIIIa), cloned from human placenta cDNA library, was overexpressed in the methylotrophic yeast Pichia pastoris GS115 (his4) and secreted into the culture medium to yield the recombinant pro-FXIIIa subunit with a predicted molecular mass of approximately 83 kDa. The gene was located immediately downstream of the strong yeast alcohol oxidase promoter (AOX1). In shake flask culture, recombinant pro-FXIIIa (rFXIIIa) was secreted into the culture medium at above 50 mg l^–1. The fibrin-stabilizing activity of the recombinant pro-FXIIIa, after thrombin activation, was confirmed using fibrin cross-linking patterns, and analyzed by SDS-PAGE.     

Application of the Fröhlich theory to the modelling of rouleau formation in human erythrocytes

The details of Fröhlich's theory and some recent experiments on the rouleau formation of human erythrocytes which exhibit a strong interaction that appears to satisfy the prerequisites of the Fröhlich theory, are summarized. To verify whether the Fröhlich theory of long-range coherence in biological systems is applicable to the phenomenon of rouleau formation in human erythrocytes, the interactions between erythrocytes are modelled as those between two large, coupled oscillating dipoles. Relevant expressions for the resonant long-range and the van der Waals interaction are then derived. Using the available numerical data, the eigenfrequencies and the interaction energies corresponding to the experimental conditions are then derived. In the range of postulated frequencies (10¹¹–10¹² Hz) the effective interaction coefficient due to the resonant long-range forces is, indeed, found to agree with its experimental value of 3.0. However, the same value of can also be achieved through the ordinary van der Waals interactions between dipoles oscillating at lower frequencies. It is concluded that the resonant long-range interaction between erythrocytes may be responsible for the onset of rouleau formation. However, other mechanisms cannot be ruled out at this stage, especially since the Fröhlich mechanism requires a number of unconfirmed preconditions.     

Contribution of α-adrenoceptors to the contractility of the human myocardium in chronic coronary heart disease

The inotropic response of isolated myocardial strips to ₁-adrenoceptor stimulation was compared for patients with chronic coronary heart disease (CHD) and patients with WPW syndrome. The ₁-adrenoceptors were stimulated with 1 × 10^– M phenylephrine after blocking of the -adrenoceptors with 3 × 10^–1 M propranolol. The inotropic activity was recorded in the isometric mode. In the myocardium without signs of ischemic damage, stimulation of the ₁-receptors caused a slowly developing single-phase positive inotropic response. The myocardium of the CHD patients was characterized by a three-phase response. The specific features of the inotropic response to ₁-adrenoceptor stimulation in the CHD patients were assumed to be determined by changes in intracellular homeostasis of Ca²⁺. Electromechanical coupling in cardiac myocytes of CHD patients depends on Ca²⁺ deposited in the sarcoplasmic reticulum to a greater extent than coupling in the intact myocardium. An additional positive inotropic effect is possible upon exogenous calcium influx into cardiac myocytes.Translated from Fiziologiya Cheloveka, Vol. 31, No. 1, 2005, pp. 133–136.Original Russian Text Copyright © 2005 by Afanasev, Ugdyzhekova, Karpov.     

Is the development of falciparum malaria in the human host limited by the availability of uninfected erythrocytes?

Background  

The development and propagation of malaria parasites in their vertebrate host is a complex process in which various host and parasite factors are involved. Sometimes the evolution of parasitaemia seems to be quelled by parasite load. In order to understand the typical dynamics of evolution of parasitaemia, various mathematical models have been developed. The basic premise ingrained in most models is that the availability of uninfected red blood cells (RBC) in which the parasite develops is a limiting factor in the propagation of the parasite population.