Circadian and circannual study of the hypophyseal-gonadal axis in healthy young males

Circadian and circannual variations of Testosterone, FSH and LH secretions, other than Oral Body Temperature (OBT) have been studied in four healthy males. OBT showed a constant circadian rhythm with an acrophase located in the afternoon. Plasma Testosterone exhibited both a circadian (acrophase = hr 09,28) and a circannual rhythm (acrophase = 22 february); plasma FSH also showed a circannual rhythm (acrophase = 13 february). By mean chronogram +/- SEM we documented the highest LH levels in December and the lowest in February. These observations would suggest the hypothesis that the winter could be the period in which the hypophysis-gonadal axis in young males exhibits its maximal activity as previously documented for other hormones.     

Identification and characterization of rDJL, a novel member of the DnaJ protein family, in rat testis

Applying the method of segmentation of seminiferous tubules combined with DDRT-PCR and cDNA library screening, a novel DnaJ homologue, rDJL was identified in rat testis. The reading frame encodes a protein of 223 amino acid residues containing J domain in the NH2 terminal region. rDJL gene is expressed mainly in testis and rDJL protein was immunolocalized notably in the acrosome region of spermatozoa. Immunoprecipitation experiments showed that rDJL interacted with Hsc70 and clathrin protein. When CHO cells were treated with EGF, rDJL and clathrin protein were found to be colocalized and be concentrated as endosome vesicles. The present findings suggest that rDJL functions as co-chaperone to Hsc70, participates in vesicular trafficking and may play an important role in acrosomogenesis.     

Prodigiosins: a novel family of immunosuppressants with anti-cancer activity

Prodigiosins (PrGs) are a family of promising therapeutic molecules, isolated mostly from Gram-negative bacteria and characterized by a common pyrryldipyrrylmethene structure with varying side chains. They show a broad spectrum of activities such as anti-microbial, anti-malarial, anti-cancer and immunosuppressive. PrGs are attracting increasing attention due to the ongoing research for less toxic, but effective agents for cancer chemotherapy and immunosuppression for preventing allograft rejection and autoimmunity. Different analogues have been synthesized and evaluated. This review discusses the immunosuppressive and anti-cancer activities of this class of compounds, as both involve inhibition of cell proliferation. The main focus is on the in vitro and in vivo immunosuppressive activity of the different PrGs and the mechanisms involved. PrGs primarily target the T cells, though some effects are observed on other cell types also. Unlike the well-known immunosuppressant cyclosporin A, PrGs do not inhibit the secretion of IL-2 but inhibit the mitogenic signaling from IL-2, suggesting a different mechanism of action. Janus tyrosine kinase 3 (Jak3) that associates with IL-2R upon activation is considered as the molecular target for PrGs. Its restricted expression makes Jak3 as an attractive target for immunosuppressive therapy. However, the available literature suggests that some other pathways are also influenced by the PrGs. These may be important for the anti-cancer activity, as well as immunosuppressive action. Therefore, PrGs appear to be potential candidates for pharmaceutical development as immunosuppressants and also as anti-cancer agents.     

Synthesis,in vitro antimalarial activity and cytotoxicity of novel 4-aminoquinolinyl-chalcone amides

A series of 4-aminoquinolinyl-chalcone amides 11–19 were synthesized through condensation of carboxylic acid-functionalized chalcone with aminoquinolines, using 1,1′-carbonyldiimidazole as coupling agent. These compounds were screened against the chloroquine sensitive (3D7) and chloroquine resistant (W2) strains of Plasmodium falciparum. Their cytotoxicity towards the WI-38 cell line of normal human fetal lung fibroblast was determined. All compounds were found active, with IC₅₀ values ranging between 0.04–0.5 μM and 0.07–1.8 μM against 3D7 and W2, respectively. They demonstrated moderate to high selective activity towards the parasitic cells in the presence of mammalian cells. However, amide 15, featuring the 1,6-diaminohexane linker, despite possessing predicted unfavourable aqueous solubility and absorption properties, was the most active of all the amides tested. It was found to be as potent as CQ against 3D7, while it displayed a two-fold higher activity than CQ against the W2 strain, with good selective antimalarial activity (SI = 435) towards the parasitic cells. During this study, amide 15 was thus identified as the best drug-candidate to for further investigation as a potential drug in search for new, safe and effective antimalarial drugs.     

Periventricular and suprachiasmatic lesion effects on photoperiodic responses of the hamster hypophyseal-gonadal axis

We examined the involvement of neural mechanisms within the suprachiasmatic nucleus (SCN) and periventricular area (PVA), and the role of prolactin (Prl) in control of endocrine function in short day-exposed Syrian hamsters. Hamsters bearing lesions of the SCN or PVA, hamsters implanted with an anterior pituitary under the kidney capsule to provide sustained Prl levels, and sham-operated hamsters were exposed to either 14L:10D or 8L:16D. After 9 wk, hamsters were sacrificed, and their testes and pituitaries were studied in vitro to assess their secretory capacity. SCN lesions and large periventricular lesions impinging on the paraventricular nucleus prevented testicular regression during short-day exposure. Small periventricular lesions and pituitary implants did not prevent gonadal regression in hamsters exposed to short days. Testis weights were positively correlated with basal and luteinizing hormone (LH)-stimulated androgen production in the control and lesioned groups; pituitary implants prevented the decline in androgen production in vitro in gonadally regressed animals. The relative in vitro pituitary response to gonadotropin-releasing hormone (GnRH) stimulation in control and lesioned groups was not reduced by short-day exposure. These data indicate that either axons coursing dorsally from the SCN or extra-SCN structures in the periventricular/paraventricular area are necessary for testicular regression in short photoperiods.     

Piperidyl amides as novel,potent and orally active mGlu5 receptor antagonists with anxiolytic-like activity

High throughput screening led to the identification of nicotinamide derivative 2 as a structurally novel mGluR5 antagonist. Optimization of the modular scaffold led to the discovery of 16m, a compound with high affinity for mGluR5 and excellent selectivity over other glutamate receptors. Compound 16m exhibits a favorable PK profile in rats, robust anxiolytic-like effects in three different animal models of fear and anxiety, as well as a good PK/PD correlation.     

NYD-SP16, a novel gene associated with spermatogenesis of human testis

By hybridizing human adult testis cDNA microarrays with human adult and embryo testis cDNA probes, a novel human testis gene NYD-SP16 was identified. NYD-SP16 expression was 6.44-fold higher in adult testis than in fetal testis. NYD-SP16 contains 1595 base pairs (bp) and a 762-bp open reading frame encoding a 254-amino acid protein with 73% amino acid sequence identity with the mouse testis homologous protein. The NYD-SP16 gene was localized to human chromosome 5q14. The deduced structure of the NYD-SP16 protein contains one transmembrane domain, which was confirmed by GFP/NYD-SP16 fusion protein expression in the cytomembrane of the transfected human choriocarcinoma JAR cells, suggesting that it is a transmembrane protein. Multiple tissue distribution indicated that NYD-SP16 mRNA is highly expressed in the testes and pancreas, with little or no expression elsewhere. Further analysis of abnormal expression in infertile male patients revealed complete absence of NYD-SP16 in the testes of patients with Sertoli-cell-only syndrome and variable expression in patients with spermatogenic arrest. Homologous gene expression in mouse testis was confirmed in spermatogenic cells by in situ hybridization. The results of cDNA microarray, in situ hybridization, and semiquantitative polymerase chain reaction in mouse testis of different stages indicated that NYD-SP16 expression is developmentally regulated. These results suggest that the putative NYD-SP16 protein may play an important role in testicular development/spermatogenesis and may be an important factor in male infertility.     

Identification of a novel gene family,paralogs of inhibitor of apoptosis proteins present in plants,fungi, and animals

Only few orthologs of animal apoptosis regulators have been found in plants. Recently, the ectopic expression of mammalian inhibitor of apoptosis proteins (IAPs) has been shown to affect plant programmed cell death. Here, we identified two novel proteins homologous to Arabidopsis thaliana IAP-like protein (AtILP) 1 and 2 by applying an improved motif searching method. Furthermore, homologs of AtILP1 were found to occur as a novel gene family in other organisms such as fungi and animals including Homo sapiens (HsILP1). Like baculovirus IAP repeats (BIRs) in IAPs, ILPs contain two highly conserved BIR-like domains (BLDs) with a putative C2HC-type zinc finger. Phylogenetic analyses indicated that ILPs are putative paralogs of IAPs. Homology modeling revealed that the three-dimensional structure of BLD in HsILP1 is similar to that of BIR. Transient expression of HsILP1 resulted in inhibition of etoposide-induced apoptosis in HEK293 and HeLaS3 cells. These findings suggest that ILPs are conserved in a wide range of eukaryotes including plants, and that their functions are closely related to those of IAPs.     

Synthesis and study of anti-inflammatory activity of some novel cyclophane amides

Macrocyclic di- and tetra-amides with thia- and oxylinkages were synthesized and screened for in vitro anti-inflammatory activity. Cyclophane diamide 15 showed a dose-dependent activity, while the other cyclophane amides 16-20 exhibited mild activity.     

Nocathiacin analogs: Synthesis and antibacterial activity of novel water-soluble amides

Novel water-soluble amide analogs were synthesized from nocathiacin I (1) through the formation of the carboxylic acid intermediate followed by coupling to primary or secondary amines. Several compounds with potent antibacterial activity and adequate water solubility were identified. Of these, compound 19 was selected for more extensive evaluation because of its excellent in vitro antibacterial activity and in vivo efficacy, as well as clean off-target screening.     

Hainanenins: a novel family of antimicrobial peptides with strong activity from Hainan cascade-frog, Amolops hainanensis

Antimicrobial peptides (AMPs) secreted by amphibian skin represent an important innate immune defense strategy. There are more than 340 species in the family of Ranidae worldwidely, and from which nearly 100 families of AMPs comprising between 8 and 48 amino acid (aa) residues have been characterized. In current work, two novel AMPs were purified from the skin secretion of Hainan cascade-frog, Amolops hainanensis, and 31 cDNA sequences encoding 10 novel AMPs belonging to 4 families were cloned from the constructed skin cDNA library of A. hainanensis. Among these 10 AMPs, 5 peptides represent the prototypes of a novel amphibian AMP family. According to the generic name of the species of origin, they were designated as hainanenin-1-5. Each of them consists of 21 aa residues with a C-terminal disulphide loop of 7 residues between Cys(15) and Cys(21). Two of them (hainanenin-1 and 5) were then synthesized and their in vitro activities were screened, including antimicrobial, hemolytic and antioxidant activities. The results showed that hainanenin-1 and 5 possessed strong and broad-spectrum antimicrobial activities against Gram-positive, Gram-negative bacteria and fungi, including a large number of clinically isolated drug-resistant pathogenic microorganisms, and slight antioxidant activity. Undesirably, hainanenin-1 and 5 exhibited strong hemolytic activity on human erythrocytes. The discovery of hainanenins and their great antimicrobial potency provides new templates for anti-infective agent design.     

A novel,evolutionarily conserved gene family with putative sequence-specific single-stranded DNA-binding activity

Complete and partial deletions of chromosome 5q are recurrent cytogenetic anomalies associated with aggressive myeloid malignancies. Earlier, we identified an approximately 1.5-Mb region of loss at 5q13.3 between the loci D5S672 and D5S620 in primary leukemic blasts. A leukemic cell line, ML3, is diploid for all of chromosome 5, except for an inversion-coupled translocation within the D5S672-D5S620 interval. Here, we report the development of a bacterial artificial chromosome (BAC) contig to define the breakpoint and the identification of a novel gene SSBP2, the target of disruption in ML3 cells. A preliminary evaluation of SSBP2 as a tumor suppressor gene in primary leukemic blasts and cell lines suggests that the remaining allele does not undergo intragenic mutations. SSBP2 is one of three members of a closely related, evolutionarily conserved, and ubiquitously expressed gene family. SSBP3 is the human ortholog of a chicken gene, CSDP, that encodes a sequence-specific single-stranded DNA-binding protein. SSBP3 localizes to chromosome 1p31.3, and the third member, SSBP4, maps to chromosome 19p13.1. Chromosomal localization and the putative single-stranded DNA-binding activity suggest that all three members of this family are capable of potential tumor suppressor activity by gene dosage or other epigenetic mechanisms.     

BSPRY, a novel protein of the Ro-Ret family

Utilizing the yeast two-hybrid system we have identified a novel protein of the Ro-Ret family that was termed BSPRY. This protein is composed of a B-box, an alpha-helical coiled coil and a SPRY domain. BSPRY from human beings shares 80% sequence identity with the homologous protein from mice. The gene for BSPRY resides on human chromosome 9 and is specifically expressed in testis. It comprises six exons and five introns and possesses a GC rich promoter forming a typical CpG island. The function of BSPRY is not known, but several related proteins of the RBCC family have been implicated in cell transformation.