Functional analysis of a subset of canine olfactory receptor genes

In this paper, we explored the level complexity of the combinatorial olfactory code that allows mammals with a repertoire of about thousand putatively active olfactory receptors encoded in their genomes to recognize and identify a much larger repertoire of odorant molecules. To that end, we cloned 38 canine OR genes belonging to the same OR gene family and transiently expressed them in a subclone of embryonic human kidney cells (HEK293) permanently expressing the G(olf) subunit. Using a Ca(2+) imaging approach, we established for example that as many as 26 out of the 38 cloned OR elicited a Ca(2+) response when exposed to octanal, whereas 10 responded to nonanal, other aldehydes providing intermediate responses. Altogether, these results demonstrated that the combinatorial code is quite complex in support to the highly developed sense of olfaction demonstrated by dogs.     

Assessment of Phytochemical Analysis,Nutritional Composition and Antimicrobial Activity of<i> Moringa oleifera</i>

Moringa oleifera is a miracle plant rich in nutrients, antioxidants, and antibiotic properties. Present study was designed to evaluate various biochemical attributes of leaves and flowers of M. oleifera. Plant parts (leaves, flowers) of M. oleifera, collected from different roadsides of Multan district, Punjab, Pakistan, were used as experimental material. Result indicates that alkaloids, saponin, carbohydrates, fats, and protein had a high value in the aqueous extract of both leaves and flowers of M. oleifera. Whereas phenol content was high in methanolic leaves extract and the phenol contents were high in aqueous extract of flowers. The extract yield of M. oleifera leaves and flowers both showed a higher percentage in aqueous extract (57.5%), followed by methanol extract and lowest in ethyl acetate extract. Flavonoids contents were higher in ethyl acetate extract of leaves (33.67%) and aqueous extract of flowers (53.71%). While crude fiber was high in methanolic extract of leaves (12.40%) and in flowers crude fiber was high in ethyl acetate extract (15.86%). The moisture contents were higher in leaves (8.87%) than flowers (7.3%) and similarly, ash percentage in flowers (52.60%) than leaves (41.84%). Ethyl acetate extracts of M. oleifera leaves show antibacterial activity against Pseudomonas aeruginosa while methanolic extract of M. oleifera flowers shows antibacterial activity against Xanthomonas sp. Maximum growth inhibits show in all extracts of leaves against Aspergillus flavus, F. oxysporum, and P. glabrum except for the concentrated aqueous extract of leaves. While in flowers maximum growth inhibits all extracts against P. glabrum, A. niger, and A. flavus except the diluted ethyl acetate extract. Phytochemicals present in different parts of moringa have significant edible and commercial potential. Moringa extracts exhibited significant antimicrobial activity, therefore have applications in pharmaceuticals.     

Kidney Dysfunction in Adult Offspring Exposed In Utero to Type 1 Diabetes Is Associated with Alterations in Genome-Wide DNA Methylation

Background

Fetal exposure to hyperglycemia impacts negatively kidney development and function.

Objective

Our objective was to determine whether fetal exposure to moderate hyperglycemia is associated with epigenetic alterations in DNA methylation in peripheral blood cells and whether those alterations are related to impaired kidney function in adult offspring.

Design

Twenty nine adult, non-diabetic offspring of mothers with type 1 diabetes (T1D) (case group) were matched with 28 offspring of T1D fathers (control group) for the study of their leukocyte genome-wide DNA methylation profile (27,578 CpG sites, Human Methylation 27 BeadChip, Illumina Infinium). In a subset of 19 cases and 18 controls, we assessed renal vascular development by measuring Glomerular Filtration Rate (GFR) and Effective Renal Plasma Flow (ERPF) at baseline and during vasodilatation produced by amino acid infusion.

Results

Globally, DNA was under-methylated in cases vs. controls. Among the 87 CpG sites differently methylated, 74 sites were less methylated and 13 sites more methylated in cases vs. controls. None of these CpG sites were located on a gene known to be directly involved in kidney development and/or function. However, the gene encoding DNA methyltransferase 1 (DNMT1)—a key enzyme involved in gene expression during early development–was under-methylated in cases. The average methylation of the 74 under-methylated sites differently correlated with GFR in cases and controls.

Conclusion

Alterations in methylation profile imprinted by the hyperglycemic milieu of T1D mothers during fetal development may impact kidney function in adult offspring. The involved pathways seem to be a nonspecific imprinting process rather than specific to kidney development or function.     

A new dendritic cell vaccine generated with interleukin-3 and interferon-β induces CD8+ T cell responses against NA17-A2 tumor peptide in melanoma patients

Dendritic cells derived from monocytes cultured in the presence of type I interferon were found to induce efficient T cell responses against tumor antigens in vitro. We vaccinated eight stage III or IV melanoma patients with dendritic cells generated with interferon-β and interleukin-3, activated by poly I: C, and pulsed with the tumor-specific antigen NA17.A2. This dendritic cell vaccine was well-tolerated with only minor and transient flu-like symptoms and inflammatory reactions at the injection sites. In most patients, isotopic imaging documented dendritic cells (DC) migration from the intradermal injection site to the draining lymph nodes. Finally, mixed lymphocyte-peptide culture under limiting dilution conditions followed by tetramer labeling indicated that three out of eight patients mounted a CD8 T cell response against the NA17.A2 antigenic peptide. We conclude that DC generated in type I-IFN represent an interesting alternative to DC generated in IL-4 and GM-CSF for cancer immunotherapy.     

Microtubule-associated STOP protein deletion triggers restricted changes in dopaminergic neurotransmission

The microtubule-associated stable tubule only polypeptide (STOP) protein plays a key-role in neuron architecture and synaptic plasticity. Recent studies suggest that schizophrenia is associated with alterations in the synaptic connectivity. Mice invalidated for the STOP gene display phenotype reminiscent of some schizophrenic-like symptoms, such as behavioral disturbances, dopamine (DA) hyper-reactivity, and possible hypoglutamatergia, partly improved by antipsychotic treatment. In the present work, we examined potential alterations in some DAergic key proteins and behaviors in STOP knockout mice. Whereas the densities of the DA transporter, the vesicular monoamine transporter and the D₁ receptor were not modified, the densities of the D₂ and D₃ receptors were decreased in some DAergic regions in mutant versus wild-type mice. Endogenous DA levels were selectively decreased in DAergic terminals areas, although the in vivo DA synthesis was diminished both in cell bodies and terminal areas. The DA uptake was decreased in accumbic synaptosomes, but not significantly altered in striatal synaptosomes. Finally, STOP knockout mice were hypersensitive to acute and subchronic locomotor effects of cocaine, although the drug equally inhibited DA uptake in mutant and wild-type mice. Altogether, these data showed that deletion of the ubiquitous STOP protein elicited restricted alterations in DAergic neurotransmission, preferentially in the meso-limbic pathway.     

Nuclease A (Gbs0661), an extracellular nuclease of Streptococcus agalactiae,attacks the neutrophil extracellular traps and is needed for full virulence

Most bacteria of the genus Streptococcus are opportunistic pathogens, and some of them produce extracellular DNases, which may be important for virulence. Genome analyses of Streptococcus agalactiae (GBS) neonate isolate NEM316 revealed the presence of seven genes putatively encoding secreted DNases, although their functions, if any, are unknown. In this study, we observed that respiration growth of GBS led to the extracellular accumulation of a putative nuclease, identified as being encoded by the gbs0661 gene. When overproduced in Lactococcus lactis, the protein was found to be a divalent cation‐requiring, pH‐stable and heat‐stable nuclease that we named Nuclease A (NucA). Substitution of the histidine¹⁴⁸ by alanine reduced nuclease activity of the GBS wild‐type strain, indicating that NucA is the major nuclease ex vivo. We determined that GBS is able to degrade the DNA matrix comprising the neutrophil extracellular trap (NET). The nucA^H148A mutant was impaired for this function, implicating NucA in the virulence of GBS. In vivo infection studies confirmed that NucA is required for full infection, as the mutant strain allowed increased bacterial clearance from lung tissue and decreased mortality in infected mice. These results show that NucA is involved in NET escape and is needed for full virulence.