In vivo measurement of intragastric pressure with a rubber balloon in the anesthetized rat.

The protocols described in this unit are designed to measure the intragastric pressure in anesthetized rats by a water-filled low-compliance rubber balloon. The balloon is introduced into the stomach either orally (by passing the balloon down the esophagus) or directly via a small incision of the fundus after laparotomy. The effects of both stimulatory (e.g., carbachol) and inhibitory (e.g., oxymetazoline) agents can be evaluated on the gastric tone and phasic contractions. The model allows the evaluation of dose-response curves and also the time-course of the effects. Furthermore, by combining centrally or peripherally acting agents the site of action can also be determined. Curr. Protoc. Toxicol. 57:21.12.1-21.12.11. (c) 2013 by John Wiley & Sons, Inc

Brain neuropeptides in gastric mucosal protection.

The centrally induced gastroprotective effect of neuropeptides has been intensively studied. Besides many similarities, however, differences can also be observed in their gastroprotective actions. The gastroprotective dose-response curve proved to be either sigmoid, or bell-shaped. Additional gastrointestinal effects of neuropeptides can contribute to their mucosal protective effect. Part of the neuropeptides induces gastroprotection by peripheral administration as well. Besides vagal nerve the sympathetic nervous system may also be involved in conveying the central effect to the periphery. Better understanding of the complex mechanism of the maintenance of gastric mucosal integrity may result in the development of new strategy to enhance gastric mucosal resistance against injury

Role of cannabinoids in gastrointestinal mucosal defense and inflammation.

Modulating the activity of the endocannabinoid system influences various gastrointestinal physiological and pathophysiological processes, and cannabinoid receptors as well as regulatory enzymes responsible for the synthesis or degradation of endocannabinoids represent potential targets to reduce the development of gastrointestinal mucosal lesions, hemorrhage and inflammation. Direct activation of CB1 receptors by plant-derived, endogenous or synthetic cannabinoids effectively reduces both gastric acid secretion and gastric motor activity, and decreases the formation of gastric mucosal lesions induced by stress, pylorus ligation, nonsteroidal anti-inflammatory drugs (NSAIDs) or alcohol, partly by peripheral, partly by central mechanisms. Similarly, indirect activation of cannabinoid receptors through elevation of endocannabinoid levels by globally acting or peripherally restricted inhibitors of their metabolizing enzymes (FAAH, MAGL) or by inhibitors of their cellular uptake reduced the gastric mucosal lesions induced by NSAIDs in a CB1 receptor-dependent fashion. Dual inhibition of FAAH and cyclooxygenase induced protection against both NSAID-induced gastrointestinal damage and intestinal inflammation. Moreover, in intestinal inflammation direct or indirect activation of CB1 and CB2 receptors exerts also multiple beneficial effects. Namely, activation of both CB receptors was shown to ameliorate intestinal inflammation in various murine colitis models, to decrease visceral hypersensitivity and abdominal pain, as well as to reduce colitis-associated hypermotility and diarrhea. In addition, CB1 receptors suppress secretory processes and also modulate intestinal epithelial barrier functions. Thus, experimental data suggest that the endocannabinoid system represents a promising target in the treatment of inflammatory bowel diseases, and this assumption is also confirmed by preliminary clinical studies

Gut inflammation: current update on pathophysiology, molecular mechanism and pharmacological treatment modalities

Inflammatory bowel disease (IBD) is a chronic and relapsing inflammatory condition of the gastrointestinal tract. The two main forms of IBD are Crohn's disease and ulcerative colitis. According to the recent concept the disease is caused by a combination of factors, including genetics, immune dysregulation, barrier dysfunction and the change in microbial flora. Environmental factors, such as changes in diet, antibiotic use, smoking or improved domestic hygiene (e.g. eradication of intestinal helminths) probably contribute to the development and increased prevalence of IBD. Dysregulation of mucosal immunity in IBD causes an overproduction of inflammatory cytokines which resulted in uncontrolled intestinal inflammation. Based on extensive research over the last decade, besides the conventional therapy, there are several novel pathways and specific targets, on which focus new therapeutics. New therapeutics aim 1./ to correct genetic susceptibility by stimulating NOD2 expression, TLR3 signaling or inhibition of TLR4 pathway, 2./ to restore the immune dysregulation by inhibition of pro-inflammatory cytokines (TNF-α, IL-6, IL-13, IL-17, IL-18, IL-21), Th1 polarisation (IL-2, IL-12, IL-23, IFN-γ ), T-cell activation, leukocyte adhesion, as well as by immunostimulation (GM-CSF, G-CSF) and anti-inflammatory cytokines (IL-10, IL-11, IFN-β-1a), 3./ to restore mucosal barrier function and stimulate mucosal healing by different growth factors, such as GH, EGF, KGF, TGF-β, VEGF, 4./ to restore the normal bacterial flora by antibiotics, probiotics. However, in spite of these numerous potential targets, the true value and clinical significance of most of the new biologics and molecules are not clear yet

Comparative in vivo analysis of recombinant type II feline coronaviruses with truncated and completed ORF3 region.

Our previous in vitro comparative study on a feline coronavirus (FCoV) pair, differing only in the intactness of their ORF3abc regions, showed that the truncated ORF3abc plays an important role in the efficient macrophage/monocyte tropism of type II feline infectious peritonitis virus (FIPV). In the present study, we describe a challenge experiment with the same recombinant FCoVs in order to gain data on the in vivo characteristics on these viruses. While parent virus FIPV DF-2 developed feline infectious peritonitis in all the infected cats, its recombinant virus PBFIPV-DF-2, differing only in seven nucleotides, proved to be surprisingly low virulent, although caused an acute febrile episode similarly to the original FIPV DF-2. PBFIPV-DF-2 infection induced significantly lower virus neutralization titers than its parent virus, and lacked the second phase of viremia and development of fatal course of the disease. The recombinant PBFIPV-DF-2-R3i with completed ORF3abc gained biological properties that differentiate between the feline enteric coronavirus (FECV) and FIPV biotypes such as intensive replication in the gut, absence of viremia and weak or no serological response. Using reverse genetic approaches our study is the first experimental proof that ORF3abc is indeed responsible for the restriction of FECV replication to the intestine in vivo

A mélyfagyasztás hatásainak vizsgálata az emberi ivarsejtekre morfológiai és funkcionális paraméterek alapján = Investigation of the effects of cryopreservation on morphological and functional parameters of human gametes

-35 normozoospermiás, 27 oligozoospermiás és 42 asthenozoospermiás férfi ondómintájához pentoxyphyllin-t adtunk, emelkedő koncentrációban. Meghatároztunk azt az ideális pentoxyphyllin koncentrációt, amely leginkább javította a minta motilitását a kiindulási értékhez képest. - A komputervezérelt fagyasztóberendezés programozásának beállításával meghatároztuk az ideális fagyasztási algoritmust. - A komputervezérelt fagyasztóberendezéssel végzett ondófagyasztást követő felolvasztás után pentoxyphyllin hozzáadásával vizsgáltuk a motilitást. Az optimális motilitásnövelő hatást 0,75 mmol/l pentoxphyllin-koncentráció esetén kaptuk. -74 donormintát fagyasztottunk le. Összesen 197 asszonyt kezeltünk a fagyasztott mintákkal, amely összesen 280 intrauterin inszeminációs ciklust jelentett. Randomizáltan 71 ciklusban használtunk pentoxyphyllin-nel kezelt mintát. A többi ciklus kontrollként szolgált. A kumulatív terhességi arány a három ciklusra vonatkoztatva 38,2% vs. 38,0%. A különbség nem szignifikáns. -Mivel petefészek fagyasztás igénye a vizsgálati időszakban összesen két alkalommal merült fel, amelyek közül csak egy alkalommal sikerült fagyasztható mintát nyerni a petefészek-szövetből, ezért az ezzel kapcsolatos vizsgálatokat nem tudtuk elvégezni. - Eddigi eredményeinket tudományos közlemények és előadások formájában ismertettük hazai és nemzetközi folyóiratokban és fórumokon. Egy PhD disszertáció készült a fenti anyag felhasználásával. | A total of 35 normozoospermic, 27 oligozoospermic and 42 asthenozoopermic semen samples were supplemented with pentoxyphyllin in increasing concentration to define the optimal pentoxyphyllin concentration, that most effective for improving the motility of the semen sample comparing to the original value. We determined the ideal freezing algorithm with the help of an automatic computerized freezer. Motility was assessed after pentoxyphyllin supplementation of the post-thaw samples. The maximal motility increase was found at 0.75 mmol/l pentoxyphyllin concentration. A total of 74 donor semen sample was frozen, and 197 women were treated with the frozen samples in 280 intrauterine insemination cycles. Randomly, pentoxyphyllin supplemented samples were used in 71 cycles, and the cycles without using pentoxyphyllin treated samples were served as control. Cumulative pregnancy rates for the three cycles were 38.2% vs. 38.0%. The difference was not significant. As demand and endorsement of ovarium tissue cryopreservation were only two times during the study period, and in one case of them the extraction of freezable sample from the ovary was unsuccessful, hence we were not able to complete this part of the study. The reached results were presented as articles in national and international journals and presentations on national and international congresses. With the use of the above mentioned results one PhD thesis has been submitted

The SAT protein of porcine parvovirus accelerates viral spreading through irreversible ER stress induction

International audienceThe SAT protein of porcine parvovirus (PPV) accumulates in the endoplasmic reticulum (ER) and SAT deletion induces "slow spreading" phenotype. The in vitro comparison of the wild type Kresse strain and its SAT(-) knockout mutant revealed that prolonged cell integrity and late viral release are responsible for the slower spreading of the SAT(-) virus. During PPV infection, regardless of the presence or absence of SATp, the expression of downstream ER stress response proteins (Xbp1 and CHOP) was induced. However, in the absence of SATp, significant differences were detected in the quantity and the localization of CHOP, suggesting a role of SATp in the induction of irreversible ER stress in infected cells. The involvement of irreversible ER stress induction in PT cell necrosis and the viral egress was confirmed by treatment of infected cells by ER stress inducing chemicals (MG132, DTT and Thapsigargin) that accelerated the egress and spreading both the wild type and the SAT(-) viruses. UV stress induction had no beneficial effect to PPV infectionunderscoring the specificity of ER stress pathways in the process. However, induction of CHOP and its nuclear translocation cannot alone be responsible for the biological effect of SAT, since nuclear CHOP could not complement the lack of SAT in a co-expression experiment.IMPORTANCE SATp is encoded in an alternative ORF of the PPV genome. Earlier we showed that SATp of the attenuated PPV-NADL-2 strain accumulates in the ER and accelerates virus release and spreading. Our present work revealed that "slow spreading" is a general feature of the SAT(-) PPV viruses and is the consequence of prolonged cell integrity. PPV infection induced ER stress in the infected cells regardless of SATp presence, as demonstrated by the morphological changes of the ER, and expression of the stress response proteins XBP1 and CHOP. However, the presence of SATp made the ER stress more severe and accelerated the cell death during infection as shown by the higher expression rate and the alteration of the localization of CHOP. The beneficial effect of irreversible ER stress on PPV spread was confirmed by the treatment of the infected cells with ER stress inducing chemicals