Addition of Cryoprotectant Significantly Alters the Epididymal Sperm Proteome

Although cryopreservation has been developed and optimized over the past decades, it causes various stresses, including cold shock, osmotic stress, and ice crystal formation, thereby reducing fertility. During cryopreservation, addition of cryoprotective agent (CPA) is crucial for protecting spermatozoa from freezing damage. However, the intrinsic toxicity and osmotic stress induced by CPA cause damage to spermatozoa. To identify the effects of CPA addition during cryopreservation, we assessed the motility (%), motion kinematics, capacitation status, and viability of epididymal spermatozoa using computer-assisted sperm analysis and Hoechst 33258/chlortetracycline fluorescence staining. Moreover, the effects of CPA addition were also demonstrated at the proteome level using two-dimensional electrophoresis. Our results demonstrated that CPA addition significantly reduced sperm motility (%), curvilinear velocity, viability (%), and non-capacitated spermatozoa, whereas straightness and acrosome-reacted spermatozoa increased significantly (p < 0.05). Ten proteins were differentially expressed (two decreased and eight increased) (>3 fold, p < 0.05) after CPA, whereas NADH dehydrogenase flavoprotein 2, f-actin-capping protein subunit beta, superoxide dismutase 2, and outer dense fiber protein 2 were associated with several important signaling pathways (p < 0.05). The present study provides a mechanistic basis for specific cryostresses and potential markers of CPA-induced stress. Therefore, these might provide information about the development of safe biomaterials for cryopreservation and basic ground for sperm cryopreservation.     

Nitrogen and phosphorus content of flow from drains fertilized with cow manure slurry

Summary The rate of flow and nitrate and phosphorus content of the water from four drained sandy and clayey plots of size 12×60 m cropped to continuous corn were determined following two annual applications of different rates (0, 260, 390 and 520 kg N/ha) of cow manure slurry. The drain flow was directly related to the rainfall and was greatly influenced by soil texture. The N losses were greater in 1972 (7.8 to 19.1 kg N/ha) than in 1971 (0.4 to 7.8 kg N/ha) because of more summer rainfall. Nitrogen and phosphorus losses were larger from the sandy plots than from the clayey plots. The manure application rates had no apparent effect on nitrogen losses in the drain water.     

Linkages between seagrass,mangrove and saltmarsh as fish habitat in the Botany Bay estuary,New South Wales

Shallow-water vegetated estuarine habitats, notably seagrass, mangrove and saltmarsh, are known to be important habitats for many species of small or juvenile fish in temperate Australia. However, the movement of fish between these habitats is poorly understood, and yet critical to the management of the estuarine fisheries resource. We installed a series of buoyant pop nets in adjacent stands of seagrass, mangrove and saltmarsh in order to determine how relative abundance of fishes varied through lunar cycles. Nets were released in all habitats at the peak of the monthly spring tide for 12 months, and in the seagrass habitat at the peak of the neap tide also. The assemblage of fish in each habitat differed during the spring tides. The seagrass assemblage differed between spring and neap tide, with the neap tide assemblage showing greater abundances of fish, particularly those species which visited the adjacent habitats when inundated during spring tides. The result supports the hypothesis that fish move from the seagrass to the adjacent mangrove and saltmarsh during spring tides, taking advantage of high abundances of zooplankton, and use seagrass as a refuge during lower tides. The restoration and preservation of mangrove and saltmarsh utility as fish habitat may in some situations be linked to the proximity of available seagrass.     

Iron and Porphyrin Trafficking in Heme Biogenesis

Iron is an essential element for diverse biological functions. In mammals, the majority of iron is enclosed within a single prosthetic group: heme. In metazoans, heme is synthesized via a highly conserved and coordinated pathway within the mitochondria. However, iron is acquired from the environment and subsequently assimilated into various cellular pathways, including heme synthesis. Both iron and heme are toxic but essential cofactors. How is iron transported from the extracellular milieu to the mitochondria? How are heme and heme intermediates coordinated with iron transport? Although recent studies have answered some questions, several pieces of this intriguing puzzle remain unsolved.     

Risk assessment of heavy metal and metalloid toxicity through a contaminated vegetable (Cucurbita maxima) from wastewater irrigated area: A case study for a site-specific risk assessment in Jhang,Pakistan

The present research was conducted in district Jhang, Pakistan, to evaluate the concentration of metals/metalloids in soil and pumpkin (Cucurbita maxima) irrigated with domestic wastewater. Data revealed that the levels of metals and metalloids in soil samples from two different sites were below the safe limits except Cd, whereas, in the vegetable, the concentrations of As, Se, Ni, Mo, Pb, Mn, and Cu were above the safe limits. The levels of 12 metals and metalloids in the soil were ranged between 0.14 to 22.76 mg/kg at site-I and 0.16 to 22.13 mg/kg at site-II. The levels of these metals in the vegetable were found 0.35 to 61.13 mg/kg at site-I and 0.31 to 53.63 mg/kg at site-II. The transfer factor at both sites was highest for As and Co. The pollution load index recorded for Se, Cu, Cd, Mo, Pb, and Co was greater than 1. The daily intake of As, Mn, and Mo was above the oral reference dose, which reflects that the intake of pumpkin is not safe for the inhabitants of the selected sites. The control measures should be taken to phytoextract heavy metals and metalloids from polluted sites so as to reduce the health risks.     

Beneficial effects of n-hexane bark extract of Onosma echioides L. on diabetic peripheral neuropathy

Onosma echioides Linn (Boraginaceae) is the most frequently used curative herb widely used for kidney obstruction, sciatic pain, and gout. The present study was designed to investigate the therapeutic effects of n-hexane bark extract of O. echioides (OE) L. root in vivo against Streptozotocin-induced diabetic neuropathy in SD rats. For in vivo activity, the experiment was categorized into five different groups (n = 5). Group-I was considered as nondiabetic/normal control (NC) treated with 0.5% carboxymethyl cellulose (CMC), Group II as diabetic control, Group-III, IV, and V served as diabetic treated with OE 50, OE 100, and pregabalin at a dose of 50, 100, and 10 mg/kg body weight, orally, respectively. Body weight, blood glucose, oral glucose tolerance test, behavioral studies (motor coordination test, thermal hyperalgesia, cold allodynia, locomotor activity, oxidative biomarkers (thio barbituric acid reactive substances [TBARS], superoxide dismutase [SOD], glutathione [GSH], and catalase), and histopathology of the sciatic nerve were performed. Treatment with OE showed a dose-dependent increase in neuroprotective activity by improving the myelination and decreasing the axonal swelling of nerve fibers. The verdicts of behavioral activities showed a remarkable effect on animals after the treatment of extract and standard drug pregabalin. In conclusion, our findings supported the traditional application of OE and explored its importance in the management of diabetic neuropathy. Additional clinical experiments may provide novel therapeutic drugs for diabetes and its complications.     

A simple enzyme-linked immunosorbent assay for sex hormone-binding globulin

A simple enzyme-linked immunosorbent assay (ELISA) for sex hormone-binding globulin (SHBG) has been developed. Polyclonal antibody raised to SHBG purified to homogeneity was employed. The ELISA, which may be performed in under 4 h, shows no cross-reactivity with other serum proteins, has a sensitivity of less than 1.2 fmol per sample, demonstrates excellent correlation with ligand-binding techniques (r = 0.996; p less than 0.0001), and has intra- and inter-assay coefficients of variation of between 5-9% and 7-11% respectively.     

Mini-exon derived RNA gene ofLeishmania donovani: structure,organization and expression

Mini-exon derived RNA is a small nuclear RNA of trypanosomatid protozoa such asLeishmania which donates its 5′-terminal 39 nucleotides to the 5’-ends of cellular messenger RNAs by trans-splicing. We have cloned a mini-exon derived RNA gene fromLeishmania donovani and studied its organization and expression. About 200 copies of the gene per haploid genome are organized as a tandem repeat on a single chromosome. The gene is transcribed as a 95-nucleotide RNA. The first 39 nucleotides of mini-exon derived RNA is also found at the 5′-terminus of a cellular mRNA (Β-tubulin), thus confirming its identity. Sequence analysis of the gene and its flanking regions showed that while classical RNA polymerase II promoter elements such as TATA and CAAT are absent from the 5′-upstream region, intragenic sequence motifs resembling RNA polymerase III promoter elements are present. The implications of this finding for mini-exon derived RNA expression are discussed.     

Microtubule-associated protein tau. A component of Alzheimer paired helical filaments

Microtubule-associated protein tau was purified from bovine brain microtubules by either (1) phosphocellulose chromatography, (2) heat treatment at pH 6.4, (3) heat treatment at pH 2.7, (4) heat treatment at pH 2.7 followed by extraction with perchloric acid and precipitation with glycerol, or (5) by precipitation with ammonium sulfate followed by extraction with perchloric acid. All of these tau preparations reacted specifically with antibodies to Alzheimer paired helical filaments. Affinity purified antibodies to tau labeled both Alzheimer neurofibrillary tangles and plaque neurites but not amyloid in Alzheimer brain tissue sections and labeled paired helical filament polypeptides on Western blots. Human brain tau and paired helical filament polypeptides co-migrated on sodium dodecyl sulfate-polyacrylamide gels. These results suggest that tau is a major component of Alzheimer paired helical filaments.