Utišani gga-miR-142-3p u pilećem embriju: ekspresija profila XPO1.

Differential expression of gga-miR-142-3p microRNA of haemopoeitic origin during immune organ development and functional stages in chicken embryos creates new opportunities for understanding its pivotal role during embryonic developmental stages. To decipher the role of gga-miR-142-3p in-ovo knockdown was carried out with LNA modified anti-miR-gga-miR-142-3p via intravenous route at developmental and functional stages of the immune organs and other organs. Bioinformatic analysis of the genes targeted by gga-miRNA-142-3p revealed that the predicted gene XPO1 conserved binding sites at 3’UTR. The target gene XPO1 was evaluated as the validated target of gga-miR-142-3p by employing qPCR SYBR green based technology, which was evidenced by its increased expression in the tissues of gga-miR-142-3p knockdown chicken embryos. Histopathological alterations in the immune organs and visceral organs indicated that the systemic knockdown of gga-miR-124-3p led to over expression of the XPO1 gene during the embryonic stages, and changed the environment of the immune organs related to structural integrity, immune response, signal transduction and migration of B and T cells during the embryonic developmental stage in the chicken embryos. The results clearly indicated that these changes could alter the postnatal development and functions of these immune organs, and may lead to development of immuno-compromised chickens.Različita ekspresija gga-miR-142-3p mikroRNA hemopoetskog podrijetla daje nove mogućnosti razumijevanja njezine ključne uloge u embrionalnom razvoju limfnih organa i funkcionalnih zbivanja u pilećem zametku. Za otkrivanje uloge gga-miR-142-3p in ovo provedeno je utišavanje zaključanom nukleinskom kiselinom što preko anti-miR-gga-miR-142-3p preinačuje razvojne i funkcionalne sposobnosti limfnih i drugih organa. Bioinformatička analiza ciljnih gena za gga-miRNA-142-3p otkriva da gen XPO1 ima konzervirana mjesta vezanja na 3’UTR. Gen XPO1 bio je potvrđen kao cilj za gga-miR-142-3p uporabom tehnologije temeljene na qPCR SYBR zelenilu, što je bilo dokazano njegovom povećanom ekspresijom u tkivima pilećih zametaka s utišanim gga-miR-142-3p. Patohistološke promjene u imunosnim i unutarnjim organima pokazuju da sustavno utišavanje gga-miR-124-3p vodi do prevelike ekspresije gena XPO1 tijekom embrionalnog razvoja. To mijenja zadaću imunosnih organa s obzirom na strukturni integritet, imunosni odgovor, prijenos poruka te migraciju B i T limfocita tijekom razvoja pilećih zametaka. Rezultati jasno naznačuju da te promjene mogu preinačiti postnatalni razvoj i funkcije imunosnih organa te mogu dovesti do razvoja imunološki oslabljenih pilića

Investigation of Immune Biomarkers Using Subcutaneous Model of M. tuberculosis Infection in BALB/c Mice: A Preliminary Report

Evaluation and screening of vaccines against tuberculosis depends on development of proper cost effective disease models along with identification of different immune markers that can be used as surrogate endpoints of protection in preclinical and clinical studies. The objective of the present study was therefore evaluation of subcutaneous model of M.tuberculosis infection along with investigation of different immune biomarkers of tuberculosis infection in BALB/c mice. Groups of mice were infected subcutaneously with two different doses : high (2×106 CFU) and low doses (2×102 CFU) of M.tuberculosis and immune markers including humoral and cellular markers were evaluated 30 days post M.tuberculosis infections. Based on results, we found that high dose of subcutaneous infection produced chronic disease with significant (p<0.001) production of immune markers of infection like IFNγ, heat shock antigens (65, 71) and antibody titres against panel of M.tuberculosis antigens (ESAT-6, CFP-10, Ag85B, 45kDa, GroES, Hsp-16) all of which correlated with high bacterial burden in lungs and spleen. To conclude high dose of subcutaneous infection produces chronic TB infection in mice and can be used as convenient alternative to aerosol models in resource limited settings. Moreover assessment of immune markers namely mycobacterial antigens and antibodies can provide us valuable insights on modulation of immune response post infection. However further investigations along with optimization of study protocols are needed to justify the outcome of present study and establish such markers as surrogate endpoints of vaccine protection in preclinical and clinical studies in futur

Effect of repeat dose of BCG vaccination on humoral response in mice model

7-10BCG is the only vaccine presently available against tuberculosis but it is estimated to prevent only 5% of the all potentially vaccine-preventable deaths due to Tuberculosis. Keeping these in view the present study has been undertaken to evaluate the efficacy of BCG and the effect of repeat dose of BCG on antimycobacterial humoral response in mouse model. To improve BCG immunogenicity, specific anti-mycobacterial immune responses (anti-BCG titre and total IgG level) were evaluated in mouse model using boost immunization protocols with the BCG vaccine. Mice induced with a repeat dose of BCG showed an increased anti mycobacterial humoral response, which gradually declined few weeks after single dose of BCG administration. The results suggest improved efficacy of BCG vaccine by giving repeat dose of BCG that can enhance the level of immunoprotection against tuberculosis as opposed to a single BCG dose