High serum free fatty acids and low leptin levels: Plausible metabolic indicators of negative energy balance in early lactating Murrah buffaloes

Lactation is a highly demanding event in mammals, including buffaloes. It modulates the partitioning of nutrients, energy utilization, and food intake of the mother to meet her own and infant's energy needs. Failure to satisfy these energy needs leads to Negative Energy Balance (NEB). Currently, the only available indirect NEB indicator is Body Condition Score (BCS). However, direct dependency of the BCS on the peak depletion of body fat causes its inefficient use in a dairy farm. Thus, to establish objective NEB indicators in buffaloes, the serum levels of biochemical (serum β-hydroxybutyrate [BHBA] and free fatty acids [FFAs]), and endocrine (Growth Hormone [GH], insulin-like growth factor1 [IGF1], Insulin, and leptin) parameters were estimated in buffaloes. Our results revealed that serum FFA levels were significantly (p < 0.05) higher in high milk yielders (HMY) than low milk yielders (LMY) and heifers (H) during the 3rd and the 4th weeks of postpartum. The serum FFA levels were also significantly (p < 0.001) higher in the postpartum buffaloes with BCS < 3 in the field conditions. Further, serum leptin levels were significantly (p < 0.05) lower in HMY than LMY during the 3rd week of postpartum. However, the BHBA, GH, IGF1, and insulin levels were not significantly different between lactating buffaloes and H. These observations indicated that the NEB condition is probably restricted to the first month of early lactation in buffaloes. In conclusion, the simultaneous higher FFA and lower leptin levels could act as direct plausible metabolic indicators of NEB in buffaloes.     

Progesterone upregulates calcitonin gene-related peptide and adrenomedullin receptor components and cyclic adenosine 3'5'-monophosphate generation in Eker rat uterine smooth muscle cell line

Calcitonin gene-related peptide (CGRP) and adrenomedullin (AM), two potent smooth-muscle relaxants, have been shown to cause uterine relaxation. Both CGRP- and AM-binding sites in the uterus increase during pregnancy and decrease at labor and postpartum. These changes in binding sites appear to be related to the changes in calcitonin receptor-like receptor (CRLR), receptor activity-modified protein 1 (RAMP1), RAMP2, and RAMP3 mRNA levels. It is not clear, however, whether the changes in the receptor components occur in the myometrial cells and whether the steroid hormones can directly alter these receptor components in the muscle cells. In addition, the mechanism of CGRP and AM signaling in the rat myometrium is not well understood. Therefore, we examined the mRNA expression of CGRP- and AM-receptor components, G protein Galphas, CGRP, and AM stimulation of cAMP and cGMP, and the effects of progesterone on these parameters in the Eker rat uterine myometrial smooth-muscle cell line (ELT3). ELT3 cells expressed CGRP- and AM-receptor components CRLR, RAMP1, RAMP2, and RAMP3. Expression of CRLR and RAMP1 mRNA increased with progesterone treatment and decreased with estradiol-17beta treatment. However, RAMP2 and RAMP3 mRNA expressions were unaltered by both progesterone and estradiol. Progesterone increased (P<0.05) Galphas expression and augmented CGRP- and AM-induced increases in cAMP levels. In uterine smooth-muscle cells, the antagonist to Galphas protein NF449 decreased basal as well as CGRP- and AM-stimulated cAMP levels. None of the cell treatments affected cyclic GMP production. Our results suggest that the progesterone-stimulated increases in CGRP and AM receptors, Galphas protein levels, and cAMP generation in the myometrial cells may be responsible for increased uterine relaxation sensitivity to CGRP and AM during pregnancy.     

Piperine analogs as potent Staphylococcus aureus NorA efflux pump inhibitors

Based on our recent findings that piperine is a potent Staphylococcus aureus NorA efflux pump inhibitor (EPI), 38 piperine analogs were synthesized and bioevaluated for their EPI activity. Twenty-five of them were found active with potentiating activity equivalent or more than known EPIs like reserpine, carsonic acid and verapamil. The inhibitory mechanism of the compounds was confirmed by efflux inhibition assay using ethidium bromide as NorA substrate. The present communication describes the synthesis, bioevaluation and structure related activity of these efflux pump inhibitors.     

Ibuprofen loading and release in amphiphilic peptide FA32 and its derivatives: a coarse-grained molecular dynamics simulation study

A molecular dynamics simulation study is reported to investigate the loading and release of ibuprofen (IBU) in amphiphilic peptide (AF)₆H₅K₁₅ (FA32) and its derivatives (F₁₂H₅K₁₅ and F₁₆H₅K₁₅). The peptides are represented by the MARTINI coarse-grained model, and a similar model is developed here for IBU. Upon the loading of IBU in FA32, quasi-spherical core/shell structured micelles are formed. IBU is predominantly located in the hydrophobic core and covered by Phe and Ala residues, while Lys is in the hydrophilic shell. With increasing concentration of IBU, the micelles become larger due to increased hydrophobic interactions. In FA32 derivatives, the loading of IBU leads to different morphologies; particularly, a well-structured nanofibre is formed in F₁₆H₅K₁₅. Upon pH change, the release of IBU from FA32 micelles is found to be slower than from F₁₆H₅K₁₅ nanofibre, suggesting the former is better in controlled release. The simulation study reveals that IBU-loaded morphology can be altered by changing the type of peptide and has a significant effect on IBU release profile. This bottom-up insight might be useful in the rational design of carriers for efficient drug loading and release.     

Techniques for integrating ‐omics data

The challenge for -omics research is to tackle the problem of fragmentation of knowledge by integrating several sources of heterogeneous information into a coherent entity. It is widely recognized that successful data integration is one of the keys to improve productivity for stored data. Through proper data integration tools and algorithms, researchers may correlate relationships that enable them to make better and faster decisions. The need for data integration is essential for present ‐omics community, because ‐omics data is currently spread world wide in wide variety of formats. These formats can be integrated and migrated across platforms through different techniques and one of the important techniques often used is XML. XML is used to provide a document markup language that is easier to learn, retrieve, store and transmit. It is semantically richer than HTML. Here, we describe bio warehousing, database federation, controlled vocabularies and highlighting the XML application to store, migrate and validate -omics data.     

Evidence for the existence of a new receptor for CGRP,which is not CRLR

Receptors for calcitonin gene-related peptide (CGRP), a neuropeptide known to be the most potent vasodilator, are abundantly expressed in cerebellum. A monoclonal antibody to cerebellar CGRP receptors specifically detects a 66 kDa protein from rat cerebellum and other rat and human tissues, but not from SK-N-MC cells which express calcitonin receptor-like receptor (CRLR), a recently described component of CGRP receptors. In contrast, mRNA expression for CRLR was abundant in SK-N-MC cells, but it was undetectable in rat cerebellum. Furthermore, the antibody could not detect any immunoreactive protein in HEK 293 cells transiently transfected with CRLR and receptor activity-modifying protein 1 (RAMP(1)) indicating the possible existence of another CGRP receptor, which does not involve CRLR. Due to the absence of biochemical or structural data on the existence of a CGRP(2) receptor and the new data provided in this paper, we suggest to identify the two CGRP receptors as CGRP-A and CGRP-B.     

Mouse PSP94 expression is prostate tissue-specific as demonstrated by a comparison of multiple antibodies against recombinant proteins

Prostate tissue-specific gene expression is crucial for driving potentially therapeutic genes to target specifically to the prostate. Prostate secretory protein of 94 amino acids (PSP94), also known as beta-MSP (microseminoprotein), is one of the three most abundant secretory proteins of the prostate gland, and is generally considered to be prostate tissue-specific. We have previously demonstrated that the expression of the rat PSP94 gene is strictly prostate tissue-specific by an antibody against a recombinant rat PSP94. In order to study prostate targeting utilizing the PSP94 gene in a mouse pre-clinical experimental model, we need to establish antibodies against mouse PSP94 to confirm if it is prostate tissue-specific as well. In this study, firstly we raised a polyclonal antibody against a recombinant glutathione-S-transferase- (GST-) mouse mature form of PSP94. However, it showed very poor immunoreactivity against prostate tissue PSP94 as tested in Western blotting experiments. Neither antibodies against rat PSP94 nor mouse PSP94 showed significant cross-reactivity. Thus a second antibody was established against a recombinant mouse mature PSP94 containing N-terminal polyhistidines, and stronger immunoreactivity against mouse prostate tissue PSP94 was identified in Western blotting experiments. Both of these antibodies showed immunohistochemical reactivity, while the latter showed stronger reactivity in IHC when tested with different fixatives. By studying tissue distribution, we demonstrated that, as with rat PSP94, mouse PSP94 is strictly prostate tissue-specific in experiments of both Western blotting and immunohistochemistry (IHC). This conclusion was also derived from a comparison among antibodies against human, rat, and mouse PSP94, showing very different immunoreactivities in Western blotting and IHC. Finally, a competitive assay between different species was performed. We demonstrated that antibodies against PSP94 from different species (human, primate, rodents) have poor cross-reactivities. These observations also indicate that the PSP94 gene is a rapidly evolving gene in all species. Results from this study have led to the possibility of utilizing PSP94 as a targeting agent specifically to the prostate in a mouse experimental model.