Role of ultrasonography in the evaluation of the infertile male

Ultrasonography is an important diagnostic tool in the evaluation of infertile male patients. Used in the appropriate patients, ultrasonography may be helpful in confirming the presence of a clinical varicocele and diagnosing ejaculatory duct obstruction. The accuracy of ultrasonography is operator-dependent and one must be aware of the limitations of this diagnostic modality. It is ideal for the treating urologist to perform his own ultrasonographic studies when possible, particularly TRUS. The interpretation of these studies may be influenced by clinical data not available or familiar to radiologists. However, one should remain cautious about the over interpretation of ultrasonographic studies and refrain from injudicious use of this safe and relatively inexpensive imaging modality. The results of these studies may lead to inappropriate surgical interventions that are expensive and involve significant risk.     

Molecular biology of the kallikrein-kinin system: from structure to function

The participation of the kallikrein-kinin system, comprising the serine proteases kallikreins, the protein substrates kininogens and the effective peptides kinins, in some pathological processes like hypertension and cardiovascular diseases is still a matter of controversy. The use of different experimental set-ups in concert with the development of potent and specific inhibitors and antagonists for the system has highlighted its importance but the results still lack conclusivity. Over the last few years, transgenic and gene-targeting technologies associated with molecular biology tools have provided specific information about the elusive role of the kallikrein-kinin system in the control of blood pressure and electrolyte homeostasis. cDNA and genomic sequences for kinin receptors B2 and B1 from different species were isolated and shown to encode G-protein-coupled receptors and the structure and pharmacology of the receptors were characterized. Transgenic animals expressing an overactive kallikrein-kinin system were established to study the cardiovascular effects of these alterations and the results of these investigations further corroborate the importance of this system in the maintenance of normal blood pressure. Knockout animals for B2 and B1 receptors are available and their analysis also points to the role of these receptors in cardiovascular regulation and inflammatory processes. In this paper the most recent and relevant genetic animal models developed for the study of the kallikrein-kinin system are reviewed, and the advances they brought to the understanding of the biological role of this system are discussed.     

Molecular approaches to the diagnosis of male infertility

We investigated the effects of aging and/or swimming training on the antioxidant enzyme system in diaphragm of mice. Young (2 months old) and old (26 months old) male mice were swimming-trained for 6 weeks (1 h/day, 5 days/week). Cu,Zn-Superoxide dismutase (Cu,Zn-SOD) activity was significantly upregulated with aging, and swimming training definitely enhanced the activity only in young mice. Neither aging nor swimming training had overt effect on Mn-SOD activity. Glutathione peroxidase activity in young mice was significantly increased after training, but not in old mice. Both of immunoreactive Cu,Zn-SOD and Mn-SOD were significantly increased with aging but were unaffected by swimming training. Consequently, physical training significantly enhanced the specific activity of Cu,Zn-SOD in young mice, but not in old mice. Meanwhile, swimming training significantly increased xanthine oxidase activity in both age groups, the extent of the increase being greater in old mice than in young mice. We concluded that the antioxidant enzyme system in mouse diaphragm trends to be upregulated with aging, but that swimming training improved the system only in young mouse diaphragm.     

The genetics and biology of Phytophthora infestans: modern approaches to a historical challenge

Apoptosis is a highly regulated form of programmed cell death defined by distinct morphological and biochemical features; it plays an important role in embryogenesis and the maintenance of cellular and tissue homeostasis in multicellular organisms. Its perturbation has been implicated in a wide range of patho-physiological stages, including autoimmune, bacterial, and viral disease, and degenerative disorders. The critical role of apoptosis in eliminating harmful or injured cells from tissues suggests its participation in inflammatory processes and in the resolution of inflammatory reactions. In this article, we briefly review the molecular mechanism of apoptosis and the role of apoptosis in inflammation.     

Molecular biology of opioid receptors: recent advances

Endogenous opioid peptides and opiates like morphine produce their pharmacological effects through the membrane bound opioid receptors. These receptors belong to a superfamily of G-protein-coupled receptors, all of which possess seven membrane-spanning regions. Structure-activity relationship studies of opioids opened up new avenues for the pharmacological characterization of the opioid receptors. As a further advancement in this direction, molecular cloning has led to the identification of three different types of opioid receptors -- OP1 (delta), OP2 (kappa) and OP3 (mu) -- thereby supporting the results of earlier pharmacological studies which postulated their existence. The three opioid receptors are highly homologous. Consequent to the development of highly specific and selective agonists and antagonists, it was proposed that the three types of opioid receptors could be further categorized into different subtypes. However, the molecular biology data generated so far do not support the presence of the various subtypes of the three well-characterized opioid receptors. Recent strides towards the advancement of our knowledge relating to the molecular biology of these receptors have been reviewed in this article.     

Molecular biology of hepatitis D virus: research and potential for application

Superinfection by hepatitis D virus (HDV) leads to acute hepatitis and causes progression to liver cirrhosis in a significant proportion of hepatitis B surface antigen (HBsAg) carriers. Current regimens (interferon) to treat hepatitis D patients has only transient but no lasting effects. New approaches are, therefore, warranted. Recently, several laboratory studies have discovered interesting properties of HDV that may become targets for antiviral chemicals. Viral replication requires the small hepatitis delta antigen (s-HDAg). The s-HDAg is a nuclear phosphoprotein. There is evidence indicating that phosphorylation is important for HDV replication. A second step of replication requires HDV-RNA self-cleavage and self-ligation. Interestingly, one group of antibiotics, the aminoglycosides, exerts strong suppression effects on HDV ribozyme activities. In the following stage of viral assembly, two post-translational modifications, namely isoprenylation of large HDAg and glycosylation of HBsAg are involved. Agents capable of blocking the two modifications should reduce viral production. These four possible targets are reviewed. For prevention, effective vaccines are not yet available. Two novel approaches are discussed. The first demonstrates the immunogenicity of a nucleic acid vaccine in mice. The second approach assembled an empty HDV particle in yeast. Advances on such laboratory investigations may provide new methods for the control of hepatitis D in the future.     

Molecular biology in the diagnosis of cardiovascular diseases]

Genes shown to affect risk factors or protective factors with respect to coronary heart disease (CHD) have been identified at the APOB, APOAI, LPA, LDLR, APOE and CETP loci. Rare mutations (e.g., in the LDLR and APOE genes) may have a major effect, whereas genes belonging to normal polymorphism have only a moderate effect. Even genes with only a slight effect can be clinically important in combination with other genes or life-style factors. There is gene to gene interaction between LDLR and APOE genes. Important risk factors determined by genes as well as by environmental factors are homocystein and fibrinogen. In addition to traditional lipid and apoprotein measurements, the levels of Lp(a) lipoprotein, fibrinogen and homocystein should be examined in connection with diagnosing CHD cases. DNA analyses are appropriate when familial hypercholesterolemia is suspected, and it is likely that the importance of mutation analyses will increase significantly in the near future.     

Molecular biology and congenital hand anomalies: from molecules and mutations to man

Potassium channels have been implicated in central roles in activity-dependent neural plasticity. The giant fiber escape pathway of Drosophila has been established as a model for analyzing habituation and its modification by memory mutations in an identified circuit. Several genes in Drosophila encoding K+ channel subunits have been characterized, permitting examination of the contributions of specific channel subunits to simple conditioning in an identified circuit that is amenable to genetic analysis. Our results show that mutations altering each of four K+ channel subunits (Sh, slo, eag, and Hk) have distinct effects on habituation at least as strong as those of dunce and rutabaga, memory mutants with defective cAMP metabolism (). Habituation, spontaneous recovery, and dishabituation of the electrically stimulated long-latency giant fiber pathway response were shown in each mutant type. Mutations of Sh (voltage-gated) and slo (Ca2+-gated) subunits enhanced and slowed habituation, respectively. However, mutations of eag and Hk subunits, which confer K+-current modulation, had even more extreme phenotypes, again enhancing and slowing habituation, respectively. In double mutants, Sh mutations moderated the strong phenotypes of eag and Hk, suggesting that their modulatory functions are best expressed in the presence of intact Sh subunits. Nonactivity-dependent responses (refractory period and latency) at two stages of the circuit were altered only in some mutants and do not account for modifications of habituation. Furthermore, failures of the long-latency response during habituation, which normally occur in labile connections in the brain, could be induced in the thoracic circuit stage in Hk mutants. Our work indicates that different K+ channel subunits play distinct roles in activity-dependent neural plasticity and thus can be incorporated along with second messenger "memory" loci to enrich the genetic analysis of learning and memory.     

Molecular biology and immunology of acetylcholine receptor in relation to myasthenia gravis

In search for the myasthenogenic sites in the molecular structure of acetylcholine receptor (AChR) alpha-subunit, the conformation-dependent B-cell epitopes, and the MHC class II-restricted, immune cofactors-modified T-cell epitopes were studied. Using the peptides synthesized corresponding to AChR amino acid sequence, the alpha 183-200 (as an antigen to raise "blocking antibody") and the alpha 70-90, alpha 125-147 and alpha 67-76 with alpha 107-116 (as antigens to raise "binding antibody") were found immunogenic in the induction of the disease in animals. Phenotypic changes in the T-cell lineages in the thymus were discussed. An impairment of excitation-contraction coupling in some of myasthenic muscles was attributed to a defect caused by antibodies raised against ryanodine receptor protein. Myasthenia gravis patients' sera containing anti-ryanodine receptor antibodies inhibited the calcium-induced release of calcium in response to caffeine in human rhabdomyosarcoma cell line. Buffalo/Mna rats with spontaneous benign thymoma showed (1) ryanodine receptor expressed in the thymic epithelial cells, (2) anti-ryanodine receptor antibodies in serum, and (3) reduced twitch and tetanic force without abnormality in synaptic transmission and muscle membrane properties. It is suggested that thymic epithelial cell and skeletal muscle share common ryanodine receptor antigen. The finding seen in this rat strain can be a counterpart of the feature reflecting an immune attack directed against a compartment of skeletal muscle in myasthenia gravis.     

Molecular biology of the human phenol sulfotransferase gene family

Cytosolic phenol sulfotransferases (PST) catalyze the sulfation/sulfonation of various phenolic agents, including catecholamines, thyroid hormones, and drugs (e.g., minoxidil and acetaminophen), which usually results in the inactivation and subsequent excretion of the compound. Our recent efforts have focused on the cloning and sequencing of the human gene family encoding the PST isozymes, and the results are summarized in this article. Multiple PST cDNA isolates have been cloned in various laboratories representing alleles of three phenol sulfotransferase gene loci termed as STP1, STP2, and STM. All three genes have been mapped precisely to a small region on human chromosome 16p12.1-p11.2 (homologous to mouse chromosome 7). The two most closely related genes, STP1 and STP2, encode isozymes of phenol-preferring PST (P-PST) and have been mapped to a single genomic cosmid clone, thus in proximity to one another. The STM gene encoding the monoamine neurotransmitter-preferring PST (M-PST) exhibits a lower level of similarity relative to STP1 and STP2. Genomic clones have been sequenced to determine the genomic organization for each of the three highly related genes. All contain seven coding exons, with conserved intron-exon boundaries. Sequencing of individual cDNA isolates from various tissues has revealed heterogeneity in the 5' nontranslated regions, likely due to tissue-specific promoter utilization (or perhaps alternative splicing). DNA and protein polymorphisms have been identified in the population and may be useful for molecular genetic studies of the variability in the metabolism of catecholamines, thyroid hormones, and phenolic drugs, and possibly neuropsychiatric or other metabolic disorders.     

Detection of Chlamydia trachomatis in semen by the polymerase chain reaction in male members of infertile couples

We found that on pulmonary auscultation, fine crackles could be induced by changing the posture from sitting to supine and/or from supine to supine with passive leg elevation in patients without obvious congestive heart failure. We named these crackles "posturally induced crackles (PIC)." To investigate the relationship between PIC and long-term prognosis after myocardial infarction, we followed up 262 patients who recovered from acute myocardial infarction for a mean period of six years. Cardiac death occurred in three of 78 PIC-negative patients and in 28 of 143 PIC-positive patients. PIC-negative patients had a significantly better long-term prognosis than PIC-positive patients according to the Kaplan-Meier survival curves for cardiac death (p < 0.01). In a multilogistic model based on 70 appropriate cases, PIC was the third most important prognosticator after recovery from myocardial infarction and the number of diseased coronary vessels and the pulmonary capillary wedge pressure ranked first and second, respectively.     

Molecular biology in radiation therapy: the potential impact of recombinant technology on clinical practice

There is good evidence to believe that the most important lesion induced in deoxyribonucleic acid (DNA) by ionizing radiation is a double strand break. Two double strand breaks may interact and rejoin in three different ways. (a) To form a dicentric, which is a lethal event, and will lead to the death of the cell. (b) To form a symmetrical translocation, which may activate an oncogene, and result in, for example, a leukemia or lymphoma. (c) To result in a deletion, which may remove or inactivate a suppressor gene and result in, for example, a solid tumor. Genes identified in mammalian cells may be conveniently grouped into four families. Genes involved in the repair of radiation damage can greatly influence radiosensitivity. Molecular checkpoint genes hold damaged cells in G2 to check for the integrity of their chromosomes before allowing them to proceed into mitosis; consequently, an inactivated checkpoint gene can also result in increased radiosensitivity. Activated oncogenes are associated with only a small proportion of human cancers, and tend to be found more commonly in leukemias and lymphomas and less frequently with solid tumors. A reciprocal translocation is the most likely mechanism by which radiation may activate an oncogene. An inactivated or deleted suppressor gene is commonly associated with a wide range of human cancers. It is becoming increasingly evident that many common cancers do not arise randomly in the population, but that subgroups of individuals are particularly susceptible. The challenge of recombinant technology is that in the near future it may well be possible to determine at birth the susceptibilities of a given individual by identifying mutations in key genes. This is the revolution and challenge we face in the treatment of cancer.     

Molecular packing in virus crystals: geometry, chemistry, and biology

Recombinant human erythropoietin (EPO) and fluorescein isothiocyanate-labelled dextran (FITC-dextran) loaded biodegradable microspheres were prepared from poly(lactide-co-glycolide) (PLG) by a modified spray-drying technique. This microencapsulation method was compared with the water-in-oil-in-water (w/o/w) double-emulsion method. As expected, microsphere morphology, particle size and particle size distribution strongly depended on the production process. The spray-drying method was found to have a number of advantages compared to the w/o/w double-emulsion technique. The content of residual dichloromethane (DCM) in the final product was significantly lower in case of the microspheres prepared by spray-drying. Concerning EPO loaded microspheres, spray-drying yielded higher encapsulation efficiencies. Although the microspheres obtained by spray-drying are subjected to intensive mechanical and thermal stress during the preparation, the amount of aggregates of EPO in PLG microspheres were not increased compared to the w/o/w technique. Depending on the manufacturing method, addition of cyclic DL-lactide dimers (referred to as monomers in the following) affected the in vitro release profiles of EPO and FITC-dextran from PLG microspheres. Using differential scanning calorimetry it was shown that these low molecular weight substances only seem to be present inside the microspheres produced by spray-drying. DL-Lactide significantly reduced the initial burst release of both EPO and FITC-dextran. While the following release period of EPO was not affected by the DL-lactide content, a more linear FITC-dextran release pattern could be achieved. It can be concluded that the spray-drying technique provides a number of advantages compared to the w/o/w method. The modulation of protein release using low molecular weight additives is of particular interest for parenteral depot systems.