Assessment of genetic diversity of the fat-tailed Dumba sheep of India by mitochondrial and microsatellite markers.

India has a centuries-old tradition of sheep production and breeding that accomplish economic, agricultural, and religious roles. In addition to the 44 registered sheep breeds, there is a fat-tailed sheep population referred to as Dumba. This study evaluated genetic variation in Dumba sheep and its differentiation from other Indian sheep breeds using mitochondrial DNA and genomic microsatellite loci. Haplotype and nucleotide diversity based on mitochondrial DNA analysis revealed substantially high maternal genetic diversity in Dumba sheep. Major ovine haplogroups A and B observed in sheep populations across the globe registered their presence in the Dumba sheep. The molecular genetic analysis using microsatellite markers also showed high measures of allele (10.125 ± 0.762) and gene diversity (0.749 ± 0.029). Results correspond to the non-bottleneck population that is near mutation-drift equilibrium despite some deficiency in the number of heterozygotes (FIS = 0.043 ± 0.059). Phylogenetic clustering confirmed Dumba to be a distinct population. Results of this study endow authorities with critical information imperative for sustainable utilization and conservation of Indian fat-tailed sheep, which is considered to be an untapped genetic resource contributing to the food security, livelihood, and economic sustainability of rural households in marginal areas of the country.

Comparative study of functional outcomes of arthroscopic anterior cruciate ligament reconstruction using anteromedial portal and translateral all-inside technique.

Background: Anterior Cruciate Ligament (ACL) injuries are common in the active population of the Armed Forces. Symptomatic instability prompts individuals to seek a cure or a sheltered appointment. Despite the increasing numbers of ACL reconstructions performed, the outcomes have not been so spectacular with only a meager percentage of our patients returning to preinjury levels of activity. With the premise that an all-inside ACL reconstruction is likely to result in better functional outcomes, the aim of this study was to compare the short-term functional outcomes of a large consecutive series of patients undergoing ACL reconstruction using the translateral all-inside ACL reconstruction technique (AI) and standard anteromedial portal technique (AM) with a minimum follow-up of one year. Methods: A total of 240 patients with isolated ACL tear underwent ACL reconstruction via the AI or AM technique. Their preoperative and postoperative scores were compared to look for any significant differences in functional outcomes. Results: The two groups were matched for age, BMI, mechanism of injury, and interval from injury to surgery. There was no difference in their preoperative scores. Postoperatively, although there were significant improvements across both groups, there was no significant difference between the groups at any point of time. Conclusion: The AI technique has garnered interest in recent literature in addressing ACL injuries. This study found no discernible benefit of the AI technique when compared to the AM technique in terms of functionality following an ACL reconstruction at any point of time up to 1 year following surgery.

Molecules involved in the development of Barrett's esophagus phenotype in chronic eosinophilic esophagitis.

This paper aims to investigate the molecules involved in development of Barrett's esophagus (BE) in human eosinophilic esophagitis (EoE). Histopathological, immunohistochemical, real-time PCR Immuno blot, and ELISA analyses are performed to identify the signature genes and proteins involved in the progression of BE in EoE. We detected characteristic features of BE like intermediate columnar-type epithelial cells, induced BE signature genes like ErbB3, CDX1, ErbB2IP in the esophageal mucosa of patients with EoE. In addition, we had observed several BE-associated proteins such as TFF3, p53 and the progression markers like EGFR, p16, MICA, MICB, and MHC molecules in esophageal biopsies of patients with chronic EoE. Interestingly, we also detected mucin-producing columnar cells and MUC-2, MUC-4, and MUC5AC genes and proteins along with induced IL-9 in patients with chronic EoE. A strong correlation of IL-9 with mucin genes is observed that implicated a possible role for IL-9 in the transformation of esophageal squamous epithelial cells to columnar epithelial cells in patients with EoE. These findings indicate that IL-9 may have an important role in BE development in patients with chronic EoE. We also discovered that IL-9 stimulates mucin-producing and barrier cell transcripts and proteins such CK8/18, GATA4, SOX9, TFF1, MUC5AC, and tight junction proteins in primary esophageal epithelial cells when exposed to IL-9. Taken together, these findings provide evidence that indeed IL-9 has a role in the initiation and progression of BE characteristics like development of mucin-producing columnar epithelial cells in patients with chronic EoE.NEW & NOTEWORTHY Intermediate columnar-type epithelial cells are observed in biopsies of patients with EoE. Induced BE signature genes (CK8/18, CDX1 GATA4, SOX9, and Occludin) were observed in patients with chronic EoE. Induction of IL-9 and its correlation with eosinophils mucin-producing genes and proteins was observed in patients with EoE. Induced IL-9 may be responsible for the development of BE in patients with chronic EoE.

Role of IL-18-transformed CD274-expressing eosinophils in promoting airway obstruction in experimental asthma.

BACKGROUND: IL-5-dependent residential and IL-18-transformed pathogenic eosinophils have been reported; however, the role of IL-18-transformed CD274-expressing pathogenic eosinophils compared to IL-5-generated eosinophils in promoting airway obstruction in asthma has not yet been examined. METHODS: Eosinophils are detected by tissue anti-MBP and anti-EPX immunostaining, CD274 expression by flow cytometry, and airway resistance using the Buxco FinePointe RC system. RESULTS: We show that A. fumigatus-challenged wild-type mice, and different gene-deficient mice including naïve CC10-IL-18-transgenic mice, accumulate mostly peribronchial and perivascular CD274-expressing eosinophils except naïve CD2-IL-5-transgenic mice. Additionally, we show that CD2-IL-5 transgenic mice following rIL-18 treatment accumulate high number of CD274-expressing perivascular and peribronchial eosinophils with induced collagen, goblet cell hyperplasia and airway resistance compared to saline-challenged CD2-IL5 transgenic mice. Furthermore, we also show that even A. fumigatus-challenged IL-5 -/- mice and rIL-18 given ΔdblGATA mice accumulate CD274-expressing eosinophil-associated asthma pathogenesis including airway obstruction. Most importantly, we provide evidence that neutralization of CD274 and IL-18 in A. fumigatus-challenged mice ameliorate experimental asthma. Taken together, the data presented are clinically significant in establishing that anti-IL-18 neutralization is a novel immunotherapy to restrict asthma pathogenesis. CONCLUSIONS: We demonstrate that IL-18 is critical for inducing asthma pathogenesis, and neutralization of CD274 is a potential immunotherapeutic strategy for asthma.

Occurrence, molecular characterization and antimicrobial-resistance pattern of Staphylococcus species isolates from buck semen.

Staphylococcus aureus is one of the most prevalent pathogens, and a causative agent of a variety of infections in humans and animals. Most studies concentrated on characterization of staphylococcus isolates and its antimicrobial resistance from various illness of veterinary importance, but there is no specific study that is available on isolates from reproductive tract of small ruminants and especially its semen. Hence, in the current study, a total of 48 semen samples were collected from healthy bucks of different breeds to investigate the occurrence of S. aureus. Antimicrobial resistance and virulence of the Staphylococcus isolates were determined to assess the adverse effects of them on buck fertility. The bacterial isolates were tentatively confirmed as Staphylococcus spp. based on the Gram's staining, growth on Mannitol salt agar and catalase test. Overall, 75% (n = 36) of the samples were positive for Staphylococcus spp. from the total 48 buck semen ejaculates from different breeds and among them 23 (63.89%) were coagulase-negative (CoNS) and 13 (36.11%) were coagulase-positive Staphylococcus (CoPS) strains. The species identified by molecular characterization are S. aureus, S. chromogenes, S. haemolyticus, S. sciuri, S. simulans, and S. epidermidis from buck semen. Further, these isolates exhibited varying degrees of multidrug resistance genotypically as well as phenotypically. The presence of antibiotic resistance and virulence genes may pose a potential threat to reproductive health of animals, the animal handlers and livestock keepers, while simultaneously highlighting the need for vigilant monitoring of these isolates at the time of semen cryopreservation.

Hydraulic performance, consolidation characteristics and shear strength analysis of bentonites in the presence of fly-ash, sewage sludge and paper-mill leachates for landfill application.

A rapid upsurge in urban and industrial developments leads to increased generations of solid wastes. The most accepted technique of waste discarding around the world is landfilling. Leaching chemicals from municipal dumping grounds can pollute the groundwater source and the surrounding environment without appropriate precautionary measures. Bentonite is a low-cost constituent used as a liner material in landfills due to its low permeability, high sealing ability, high specific surface area, and the ability to hold up the impurity migration through adsorption. However, leachate interaction with bentonite may alter its properties and reduce its usefulness as a barrier material in the long term. Also, bentonite having different chemical and mineralogical compositions will behave differently due to the leachate interaction. Therefore, it is necessary to compare the performance of various bentonites in the presence of leachates. In the present investigation, two Indian bentonites of different mineralogical compositions were studied for their change in the index properties, swelling, swelling potential, swelling pressure, hydraulic conductivity, consolidation parameters and shear strength properties in the presence of fly ash, sewage sludge and paper mill leachates. The outcomes showed that in the presence of all the leachates, liquid limit, free swell, compression index, swelling potential, swelling pressure, time to complete 90% of consolidation and shear strength dropped; whereas, hydraulic conductivity and coefficient of consolidation increased. Besides, the quality of bentonite prominently influenced the hydraulic, strength and swelling behaviour. The bentonite having a higher cation exchange capacity, liquid limit, specific surface area, and swelling capability undergoes a higher variability in the free swell (80.0, 73.8 and 76.9% decline), liquid limit (73.5, 61.7 and 69.2% decline), swelling potential (61.3, 55.7 and 51.0% decline), swelling pressure (53.3 and 56.4% decrease), and hydraulic conductivity (57.5, 8.6 and 41.1 times increase at a void ratio of 1.2) values when infused with fly ash, sewage sludge and paper mill leachates, respectively. The study also showed that the fly ash leachate interaction causes a higher variation in bentonite behaviour than sewage sludge and paper mill leachates. The study's findings would prove beneficial to design engineers for selecting bentonite types for landfill liners.

Tacrolimus (FK506) treatment protects allergen-, IL-5- and IL-13-induced mucosal eosinophilia.

Eosinophils are a common clinical feature associated with chronic allergic diseases, and elemental diets, systemic steroids, anti-IL-5 and anti-IL-13 treatment have shown some therapeutic promise. Herein, we present evidence that pre- and post-intraperitoneal administration of tacrolimus (FK506) is very effective in reducing CCR3/Siglec-F+ eosinophils in Aspergillus-challenged asthma and EoE, CD2-IL-5 induced global eosinophilia, and DOX regulated IL-13-induced asthma. We used flow cytometry and anti-major basic protein (MBP) immunostaining to examine eosinophils in the spleen, bone marrow, BALF, lung, oesophagus and intestine. Additionally, we also performed ELISA and Western blot analyses to show that tacrolimus treatment also reduces the levels of eosinophil-specific cytokines IL-4, IL-5, IL-13 and TGF-ß, eosinophil-specific chemokines Eotaxin-1 and Eotaxin-2, and progenitors of target RCAN1 mRNA and protein levels. Additionally, the current investigations also show that the TGF-ß-mediated oesophageal and lung fibrosis is also reduced in Aspergillus-challenged, CD2-IL-5 transgenic and DOX-responsive IL-13 mice. Mechanistically, we show that tacrolimus in vitro treatment inhibited bone marrow-derived eosinophil proliferation and viability by promoting eosinophil apoptosis that may be associated with downregulation of RCAN1. Taken together, we provide in vivo and in vitro evidence that tacrolimus ameliorates eosinophil levels and associated pathogenesis in allergen-, IL-5- and IL-13-induced EoE, EG and asthma pathogenesis. Considering tacrolimus side-effects and reactivity to several other drugs, we propose the topical use of tacrolimus for paediatric and low-dose oral for adult patients as a novel therapeutic strategy for the clinical trial to reduce mucosal eosinophilia first in steroid-refractory or elemental diet non-responsive adult EoE, EG and asthma patients.

Blood mRNA levels of T cells and IgE receptors are novel non-invasive biomarkers for eosinophilic esophagitis (EoE).

Eosinophilic esophagitis (EoE) is often misdiagnosed as GERD; therefore, the goal of the current study is to establish a non-invasive diagnostic and monitoring biomarker that differentiated GERD from EoE. Reports indicates that IL-15 responsive iNKT cells and tissue specific IgE have a critical in EoE pathogenesis, not in GERD. Therefore, we tested the hypothesis that the panel of IL-15-responsive T cell and IgE receptors may be novel non-invasive biomarkers for EoE. Accordingly, the receptors of IL-15 responsive T cells (Vα24, Jα18, γδT, αßT) and IgE (FcεRI & FcεRII) were examined. The data indicates that blood mRNA levels of Vα24, Jα18, γδ T, αß T and FcεRI are significantly reduced in EoE compared to the GERD patients and normal individuals. The ROC curve analysis indicated FcεRII, Jα18 and δ TCR are the positive predictors that discriminate EoE from GERD. Thus, these molecules will be a novel non-invasive diagnostic biomarker for EoE.

5-HT1A targeting PARCEST agent DO3AM-MPP with potential for receptor imaging: Synthesis, physico-chemical and MR studies.

Contrast enhancement in MRI using magnetization or saturation transfer techniques promises better sensitivity, and faster acquisition compared to T1 or T2 contrast. This work reports the synthesis and evaluation of 5-HT1A targeted PARACEST MRI contrast agent using 1,4,7,10-tetraazacycloDOdecane-4,7,10-triacetAMide (DO3AM) as the bifunctional chelator, and 5-HT1A-antagonist methoxyphenyl piperazine (MPP) as a targeting unit. The multi-step synthesis led to the MPP conjugated DO3AM with 60% yield. CEST-related physicochemical parameters were evaluated after loading DO3AM-MPP with paramagnetic MRI active lanthanides: Gadolinium (Gd-DO3AM-MPP) and Europium (Eu-DO3AM-MPP). Luminescence lifetime measurements with Eu-DO3AM-MPP and computational DFT studies using Gd-DO3AM-MPP revealed the coordination of one water molecule (q = 1.43) with metal-water distance (rM-H2O) of 2.7 Å and water residence time (τm) of 0.23 ms. The dissociation constant of Kd 62 ± 0.02 pM as evaluated from fluorescence quenching of 5-HT1A (protein) and docking score of -4.81 in theoretical evaluation reflect the binding potential of the complex Gd-DO3AM-MPP with the receptor 5-HT1A. Insights of the docked pose reflect the importance of NH2 (amide) and aromatic ring in Gd-DO3AM-MPP while interacting with Ser 374 and Phe 370 in the antagonist binding pocket of 5-HT1A. Gd-DO3AM-MPP shows longitudinal relaxivity 5.85 mM-1s-1 with a water residence lifetime of 0.93 ms in hippocampal homogenate containing 5-HT1A. The potentiometric titration of DO3AM-MPP showed strong selectivity for Gd3+ over physiological metal ions such as Zn2+ and Cu2+. The in vitro and in vivo studies confirmed the minimal cytotoxicity and presential binding of Gd-DO3AM-MPP with 5-HT1A receptor in the hippocampus region of the mice. Summarizing, the complex Gd-DO3AM-MPP can have a potential for CEST imaging of 5-HT1A receptors.

Synthesis and evaluation of technetium-99m labelled 1-(2-methoxyphenyl)piperazine derivative for single photon emission computed tomography imaging for targeting 5-HT1A.

Quantitative changes in expression level of 5HT1A are somewhere related to common neurological disorders such as anxiety, major depression and schizophrenia. We have designed EDTA conjugated SPECT imaging probe for localization of 5HT1A receptor in brain. For designing SPECT probe we have employed the concept of bivalent approach and a homodimeric system with desirable pharmacokinetics of 5HT1A imaging. 99mTc-EDHT was also evaluated for its stability through serum stability assay and glutathione challenge experiment. Biodistribution study showed the highest accumulation of radioactivity in kidney which depicted the renal mode of excretion from the body. However in brain the uptake of 1.21% ID per gram was observed in initial 5 min of drug administration. On blocking the receptor this percent get decreased to 0.97% ID per gram. The regional distribution in brain was also performed which showed the accumulation of drug in cerebellum, cortex and hippocampus part, which are already known for 5HT1A expression. Dynamic study in rabbit is also in support of results derived from biodistribution and blood kinetics experiment. These finding suggest that 99mTc-EDHT holds promising place for further optimization before nuclear medicine applications in different animal species.

Synthesis and biological evaluation of modified laminin peptide (N2S2-KDP) with enhanced affinity for neuronal growth and targeted molecular imaging (SPECT).

An analog of γ1 laminin (RDIAEIIKDI) decapeptide has been used to augment neuronal survival and regeneration after injuries, during aging and other CNS disorder. As a prime synthetic peptide, KDI, is responsible for the neurite outgrowth of human embryonic neurons. In this study, we have designed, modified a KDI derivative and synthesized by replacing isoleucine (I) with Pro (P) amino acid at C-terminal to enhance its potency towards neurite growth. -Cys-Gly-Cys (-CGC) N2S2 motif was also incorporated in the present design for peptide radiolabeling. The modified peptide showed a better binding with the desired 3T1M receptor for neurite growth. The peptide was synthesized using solid phase peptide synthesis and Fmoc-strategy with more than 80% yield. The receptor binding studies of 99mTc-N2S2-KDP in Neuro2A cell lines showed Kd value in 31 nM range and the complex showed appreciable brain uptake in mice. The results on human SH-SY5Y indicate that the unlabeled N2S2-KDP may perhaps be useful for neurite growth in neurodegenerative disorder.

Semen quality and total microbial load: An association study in important Indian Goat breeds during different seasons.

The invasion of the male urogenital tract by microorganisms, and its subsequent effects on sperm fertilising ability, has not been well discussed in bucks. The present study was conducted to assess the bacterial load in fresh semen of the 2-6 years old bucks. For conducting the experiment, semen ejaculates from 18 bucks (6 from each breed namely Jakhrana, Jamunapari and Barbari) were used. We collected 5 ejaculates from each buck in each season (Summer-April to June, Rainy-July to Sept and Winter-November to January). Semen was collected with the artificial vagina (AV) method, and separate AV was used for each buck every time. The semen collection frequency was once in a week. Immediately after initial evaluation, collected semen samples were transferred to the microbiology laboratory of the institute. Thereafter, the semen samples were subjected to bacteriological examination to assess the microbial load. The results of the current study indicate that the microbial load in the semen was significantly (p < 0.05) higher in the Jamunapari bucks and in aged bucks. Bacteriospermia in different seasons was not significantly varied; however, nonsignificant increase in microbial load during the rainy season was observed. Overall, the average bacterial load in the semen of Jamunapari, Barbari and Jakhrana bucks was found 540.50 ± 55.88 CFU/ml, 391.81 ± 46.33CFU/ml and 388.93 ± 44.71 CFU/ml respectively. No significant difference in bacterial counts in the subsequent ejaculates among bucks was observed. Moreover, correlation analysis revealed that the proportions of motility, viability, plasma membrane integrity and acrosomal integrity were negatively influenced by the increased bacterial contamination of buck semen.

Impact of real and simulated municipal solid waste leachates on the hydraulic and swelling behaviour of bentonites for landfill application.

For the efficient functioning of a landfill, compacted bentonite is an acclaimed liner element due to its excellent adsorption capability, minimal hydraulic conductivity, and superior specific surface area (SSA). However, the leachate generation within the landfill worsens the liner material's quality, causing migration of the leachates, contaminating groundwater, and causing pollution of surrounding environment. With this perspective, a comparative assessment of the influence of real and simulated municipal solid waste (MSW) leachate on two different bentonites has been carried out in the present investigation. The two bentonites, differing precisely by their cation exchange capacity (CEC), liquid limit (LL), and swelling capability, were examined for variation in their LL, free swell (FS), and hydraulic behaviour concerning their interaction with both leachates. Results depicted that in both the leachates, LL and FS, swelling potential (SP) and pressure declined, whereas hydraulic conductivity (HC) rose. Furthermore, the bentonite quality greatly influenced the LL, FS, SP, swelling pressure, and hydraulic behaviour. Bentonite having higher CEC, SSA, and swelling ability experienced a higher variability in the LL (55.5 and 65.2% decrease), free swelling (76.9 and 83.1% decrease), SP, swelling pressure (53.3 and 56.4% decrease), and HC (13.1 and 49.4 times increase) values when permeated with simulated and real MSW leachates, respectively. The study also showed that the real MSW leachate interaction causes a higher variation in bentonite behaviour than its simulated counterpart. The study's findings would prove beneficial to design engineers for selecting bentonite types for landfill liners.

68Ga-Labeled bismacrocyclic methylene phosphonate as potential bone seeking PET radiopharmaceutical.

Phosphonates-based agents are well-known bone-seeking radiopharmaceuticals with application in detection and therapy. With higher sensitivity and resolution offered by Positron Emission Tomography (PET), tracers based on this technique are gaining huge attention. 68Ga-based generator and radiotracers render independence from the on-site cyclotron. We report the development of 68Ga-labeled DOTA-based bismacrocyclic phosphonate derivative, for bone PET imaging. The synthesis and characterization of 68Ga- DO3P-AME-DO3P was carried out in > 95% purity. The radiotracer displayed high stability and low binding affinity (<3%) to blood serum. High in vitro binding affinity were observed for synthetic hydroxyapatite, SAOS-2, osteoclast and osteoblast cells. In vivo pharmacokinetics revealed fast washout with biphasic release pattern. The deposition of radiotracer in osseous tissues was high (Bone/Muscle ratio:18), as studied from the biodistribution studies. In vivo PET/CT and biodistribution analyses revealed the ability of 68Ga-DO3P-AME-DO3P to target and accumulate in bone, thus displaying its potential as a PET bone imaging agent.

Tryptophan conjugated magnetic nanoparticles for targeting tumors overexpressing indoleamine 2,3 dioxygenase (IDO) and L-type amino acid transporter.

Tryptophan is an amino acid required by all life forms for protein synthesis and other important metabolic functions. It is metabolized in the body using the kynurenine pathway which involves the enzyme indoleamine 2,3 dioxygenase (IDO) and its transport is regulated through the L-type amino acid transporters (LAT 1). IDO and LAT 1 are found to be overexpressed in many cancers i.e., ovarian, lung colorectal etc. In this study we have used this specific interaction as the basis for designing diagnostic agent based on iron oxide nanoparticles which can specifically target the IDO/LAT 1 over expressing tumors. We have conjugated tryptophan to the surface of super-paramagnetic nanoparticles chemically using 3-aminopropyltrimethoxysilane as a linker. The synthesized tryptophan conjugated magnetic nano-conjugate has been characterized using FTIR, UV-Vis, TEM for its shape size, charge and NMR and Mass for conjugation. The magnetization studies show decrease in the magnetic behavior after conjugation however the desired super-paramagnetic property is still retained as shown by the signature sigmoidal B-H curve. The nano-conjugate shows minimal cytotoxicity over 24 h as shown by the SRB assay in two cell lines A-549, MCF-7. Using 99mTc labeling the biodistribution and the blood kinetics of the magnetic nano-conjugate was evaluated. The study highlights the suitability of the designed magnetic Nano bioconjugate as a potential bimodal diagnostic agent.

Molecular detection of important abortion-causing microorganisms in preputial swab of breeding bucks using PCR-based assays.

Infectious diseases and aetiological agents related to female reproductive systems were extensively covered compared to its male counterpart. There needs a proper study to bridge this gap, where microflora and infectious agents of both male and female reproductive are mutually intelligible. With this study, we aimed to evaluate the microbial contamination of the preputial cavity and also screened for abortion-causing agents which are zoonotic as well. In goats, such types of abortions are caused by Brucella melitensis, Chlamydophila, Campylobacter and Coxiella etc. One of the major sources of contamination of semen is the preputial cavity, which is exposed to the external environment leading to spread of infection into the female via semen straws or by natural service. In the current study, good quality bucks (n = 32, Barbari = 12, Jamunapari = 10, Jakhrana = 10) which were routinely used for semen collection were screened for their preputial swabs, for the presence of the above pathogens. For detection of Brucella melitensis, OMP31 based TaqMan® probe real-time PCR assay was used, and for Chlamydia, 16srRNA gene based SYBR® green real-time PCR assay was employed for detection of Chlamydophila abortus. While for Campylobacter spp. and Coxiella burnetii, 16srRNA gene based conventional PCR and Trans-PCR were used, respectively. In the current study, of the screened preputial swabs, none of them showed positive for Brucella and Coxiella, but of the screened 32 samples 17 showed positive for Chlamydia (53.13%) and two (6.25%) showed positive for Campylobacter spp. The current study emphasizes on the farms and laboratories which were regularly involved in screening of brucellosis also often overlook the other potential non-brucella pathogens, causing abortions eventually incurring severe economic losses to the goat keepers.

Effect of diluent sugars on capacitation status and acrosome reaction of spermatozoa in buck semen at refrigerated temperature.

OBJECTIVE: The aim of the study was to explore the possibility of a better sugar suitable for storage of goat semen at refrigerated temperature. MATERIALS AND METHOD: For this experiment, semen was collected from eight Jakhrana bucks maintained at Jakhrana unit, ICAR-CIRG, at twice a week interval using artificial vagina. Collected semen was preliminary evaluated, and better semen samples were pooled and divided into two parts. One part of the pooled semen was diluted in egg yolk, Tris, citric acid, and fructose diluter, whereas second part was diluted in egg yolk, Tris, citric acid, and glucose diluter. Then semen samples were kept in equilibration chamber for 4 h at 5 °C after proper dilution. Both the semen samples were evaluated for viability, motility, plasma membrane integrity, sperm abnormality, lipid peroxidation, acrosomal integrity, and capacitation status at 0 h, 24 h, 48 h, and 72 h after dilution. RESULTS: Significantly (P < 0.05) higher motility was observed at 24 h in extender containing glucose as compared with extender containing fructose but motility was decreased at 48 h and 72 h. Number of capacitated sperm increased significantly (P < 0.05) and acrosomal integrity was decreased significantly (P < 0.05) at 72 h in extender containing glucose. The other parameters like viability and plasma membrane integrity were decreased significantly (P < 0.05) at 72 h and lipid peroxidation as well as sperm abnormality increased significantly (P < 0.05) in extender containing glucose. CONCLUSION: From this study, it can be concluded that fructose is better diluent sugar for refrigerated storage of buck semen.

Attenuation of Allergen-, IL-13-, and TGF-α-induced Lung Fibrosis after the Treatment of rIL-15 in Mice.

Endogenous IL-15 deficiency promotes lung fibrosis; therefore, we examined the effect of induced IL-15 in restricting the progression of lung fibrosis. Our objective in this work was to establish a novel therapeutic molecule for pulmonary fibrosis. Western blot, qPCR, and ELISA were performed on the lung tissues of IL-15-deficient mice, and recombinant IL-15 (rIL-15)-treated CC10-IL-13 and CC10-TGF-α mice, and allergen-challenged CC10-IL-15 mice were examined to establish the antifibrotic effect of IL-15 in lung fibrosis. We show that endogenous IL-15 deficiency induces baseline profibrotic cytokine and collagen accumulation in the lung, and pharmacological delivery of rIL-15 downregulates Aspergillus antigen-induced lung collagen, the profibrotic cytokines IL-13 and TGF-ß1, and α-SMA+ and FSP1+ cells in mice. To confirm that overexpression of IL-15 diminishes pulmonary fibrosis, we generated CC10-rtTA-tetO7-IL-15 transgenic mice and challenged them with Aspergillus antigen. Aspergillus antigen-challenged, doxycycline (DOX)-treated CC10-IL-15 transgenic mice exhibited decreased collagen accumulation, profibrotic cytokine (IL-13 and TGF-ß1) expression, and α-SMA+ and FSP1+ cells compared with IL-15-overexpressing mice not treated with DOX. Additionally, to establish that the antifibrotic effect of IL-15 is not limited to allergen-induced fibrosis, we showed that rIL-15 or IL-15 agonist treatment restricted pulmonary fibrosis even in CC10-IL-13 and CC10-TGF-α mice. Mechanistically, we show that T-helper cell type 17 suppressor IL-15-responsive RORγ+ T regulatory cells are induced in DOX-treated, allergen-challenged IL-15-overexpressing mice, which may be a novel pathway for restricting progression of pulmonary fibrosis. Taken together, our data establishes antifibrotic activity of IL-15 that might be a novel therapeutic molecule to combat the development of pulmonary fibrosis.

Intestinal overexpression of IL-18 promotes eosinophils-mediated allergic disorders.

Baseline eosinophils reside in the gastrointestinal tract; however, in several allergic disorders, excessive eosinophils accumulate in the blood as well in the tissues. Recently, we showed in vitro that interleukin (IL)-18 matures and transforms IL-5-generated eosinophils into the pathogenic eosinophils that are detected in human allergic diseases. To examine the role of local induction of IL-18 in promoting eosinophil-associated intestinal disorders, we generated enterocyte IL-18-overexpressing mice using the rat intestinal fatty acid-binding promoter (Fabpi) and analysed tissue IL-18 overexpression and eosinophilia by performing real-time polymerase chain reaction, Enzyme-Linked Immunosorbent Assay and anti-major basic protein immunostaining. Herein we show that Fabpi-IL-18 mice display highly induced IL-18 mRNA and protein in the jejunum. IL-18 overexpression in enterocytes promotes marked increases of eosinophils in the blood and jejunum. Our analysis shows IL-18 overexpression in the jejunum induces a specific population of CD101+  CD274+ tissue eosinophils. Additionally, we observed comparable tissue eosinophilia in IL-13-deficient-Fabpi-IL-18 mice, and reduced numbers of tissue eosinophils in eotaxin-deficient-Fabpi-IL-18 and IL-5-deficient-Fabpi-IL-18 mice compared with Fabpi-IL-18 transgenic mice. Notably, jejunum eosinophilia in IL-5-deficient-Fabpi-IL-18 mice is significantly induced compared with wild-type mice, which indicates the direct role of induced IL-18 in the tissue accumulation of eosinophils and mast cells. Furthermore, we also found that overexpression of IL-18 in the intestine promotes eosinophil-associated peanut-induced allergic responses in mice. Taken together, we provide direct in vivo evidence that induced expression of IL-18 in the enterocytes promotes eotaxin-1, IL-5 and IL-13 independent intestinal eosinophilia, which signifies the clinical relevance of induced IL-18 in eosinophil-associated gastrointestinal disorders (EGIDs) to food allergens.

Potent Acetylcholinesterase Selective and Reversible Homodimeric Agent Based on Tacrine for Theranostics.

Acetylcholinesterase (AChE) has been an important biomarker for diagnosing Alzheimer's disease (AD), due to reduction in AChE activity in post-mortem brains of AD patients. A potent, selective, and reversible homodimeric inhibitor of AChE, 5-amino- N1, N3-bis(2-(1,2,3,4-tetrahydroacridin-9-ylamino)ethyl)isophthalamide (compound 4), was synthesized by using 9-alkyl(1,2,3,4-tetrahydroacridine) pharmacophore with appended functionality. In the present work, we report the synthesis of this bivalent inhibitor of AChE. The homodimeric ligand structure was designed and studied with molecular docking tools, which revealed its high affinity and interactions with active site gorge of AChE, which includes both catalytic active site (CAS) and peripheral active site (PAS). The IC50 value of this bivalent inhibitor for AChE and BuChE were 0.54 ± 0.06 and 32.49 ± 1.2 nM, respectively, with a selectivity ratio of 60.16 toward AChE. The designed ligand also showed potent inhibitory properties on PAS activity as well as on AChE-induced amyloid aggregation with low cytotoxicity on rat hippocampal neurons. The AFM images further corroborated the Aß1-42 aggregation inhibition by compound 4 to an extent similar to bis(7)-tacrine. Moreover, the bivalent ligand was also proven to be of neurogenic potential due to its ability to induce S-phase post-treatment in rat hippocampal neuronal cells. On the basis of initial results, the agent could be further explored for its theranostic value clinically, which gives the possibility of tracing the AChE levels by molecular imaging techniques in correlation with progression of neurocognitive disorders like AD for better therapy response and patient management.