Josipa Vlainić

Cyclophosphamide induced stomach and duodenal lesions as a NO-system disturbance in rats: l-NAME, l-arginine, stable gastric pentadecapeptide BPC 157

We revealed a new point with cyclophosphamide (150xa0mg/kg/day intraperitoneally for 7xa0days): we counteracted both rat stomach and duodenal ulcers and increased NO- and MDA-levels in these tissues. As a NO-system effect, BPC 157 therapy (10xa0µg/kg, 10xa0ng/kg, intraperitoneally once a day or in drinking water, till the sacrifice) attenuated the increased NO- and MDA-levels and nullified, in rats, severe cyclophosphamide-ulcers and even stronger stomach and duodenal lesions after cyclophosphamidexa0+xa0l-NAME (5xa0mg/kg intraperitoneally once a day). l-arginine (100xa0mg/kg intraperitoneally once a day not effective alone) led l-NAME-values only to the control values (cyclophosphamidexa0+xa0l-NAMExa0+xa0l-arginine-rats). Briefly, rats were sacrificed at 24xa0h after last administration on days 1, 2, 3, or 7, and assessment included sum of longest lesions diameters (mm) in the stomach and duodenum, oxidative stress by quantifying thiobarbituric acid reactivity as malondialdehyde equivalents (MDA), NO in stomach and duodenal tissue samples using the Griess reaction. All these parameters were highly exaggerated in rats who underwent cyclophosphamide treatment. We identified high MDA-tissue values, high NO-tissue values, ulcerogenic and beneficial potential in cyclophosphamide-l-NAME-l-arginine-BPC 157 relationships. This suggests that in cyclophosphamide damaged rats, NO excessive release generated by the inducible isozyme, damages the vascular wall and other tissue cells, especially in combination with reactive oxygen intermediates, while failing endothelial production and resulting in further aggravation by l-NAME which was inhibited by l-arginine. Finally, BPC 157, due to its special relations with NO-system, may both lessen increased MDA- and NO-tissues values and counteract effects of both cyclophosphamide and l-NAME on stomach and duodenal lesions.

Stress in Gastrointestinal Tract and Stable Gastric Pentadecapeptide BPC 157. Finally, do we have a Solution?

Selyes syndrome produced by diverse nocuous agents and response to damage as such means Selyes stress triad in stress coping response to reestablish homeostasis. Logically, from the gastrointestinal tract viewpoint, such organoprotective/healing response implies the angiogenic growth factors that commonly signify the healing. Thereby, the gastric pentadecapeptide BPC 157-organoprotection (huge range of beneficial effects) signifies the Selyes stress concept/stress coping response implemented in and from gastrointestinal tract, and BPC 157 as an integrative mediator that integrates the adaptive bodily response to stress. In clinical trials without side effects, LD1 not achieved, BPC 157 healing in gastrointestinal tract, and particularly the healing of the extragastrointestinal tissues (i.e., skin/tendon/ligament/muscle/bone; nerve; cornea/ brain) were referred throughout its integrative capabilities (i.e., ulcerative colitis/multiple sclerosis model equally counteracted), native in gastrointestinal tract, stability in human gastric juice (and thereby, strong efficacy and applicability), its relevance for dopamine-system function (and thereby, counteracting effects of dopamine-system dysfunction and overfunction, centrally and peripherally (mucosa maintenance); interaction with serotonin- and GABA-system)), afforded cytoprotection/adaptive cytoprotection/organoprotection (and thereby, beneficial effects on gastric and whole intestinal tract lesions and adaptation, wounds and fistulas healing, blood vessels, somatosensory neurons, NSAIDs-side effects (including also pancreas, liver, brain lesions, and blood disturbances, prolonged bleeding, thrombocytopenia, thrombosis)). Further, we combine such gut-brain axis and the NO-system where BPC 157 counteracts complications of either L-NAME application (i.e., various lesions aggravation, hypertension) or Larginine application (i.e., hypotension, prolonged bleeding, thrombocytopenia). Also, BPC 157 particularly affects genes functions (i.e., Fos, c-Jun, Egr-1), all together suggestive for an indicative generalization. Thus, we could suggest gastric pentadecapeptide BPC 157 and BPC 157 induced-organoprotection as integrative mediator that integrates the adaptive bodily response to stress, and thereby practically applied in further therapy and in effective realization of Selyes stress response.

Stable gastric pentadecapeptide BPC 157 in the treatment of colitis and ischemia and reperfusion in rats: New insights

AIM To provide new insights in treatment of colitis and ischemia and reperfusion in rats using stable gastric pentadecapeptide BPC 157. METHODS Medication [BPC 157, L-NAME, L-arginine (alone/combined), saline] was bath at the blood deprived colon segment. During reperfusion, medication was BPC 157 or saline. We recorded (USB microscope camera) vessel presentation through next 15 min of ischemic colitis (IC-rats) or reperfusion (removed ligations) (IC + RL-rats); oxidative stress as MDA (increased (IC- and IC + RL-rats)) and NO levels (decreased (IC-rats); increased (IC + RL-rats)) in colon tissue. IC + OB-rats [IC-rats had additional colon obstruction (OB)] for 3 d (IC + OB-rats), then received BPC 157 bath. RESULTS Commonly, in colon segment (25 mm, 2 ligations on left colic artery and vein, 3 arcade vessels within ligated segment), in IC-, IC + RL-, IC + OB-rats, BPC 157 (10 μg/kg) bath (1 mL/rat) increased vessel presentation, inside/outside arcade interconnections quickly reappeared, mucosal folds were preserved and the pale areas were small and markedly reduced. BPC 157 counteracted worsening effects induced by L-NAME (5 mg) and L-arginine (100 mg). MDA- and NO-levels were normal in BPC 157 treated IC-rats and IC + RL-rats. In addition, on day 10, BPC 157-treated IC + OB-rats presented almost completely spared mucosa with very small pale areas and no gross mucosal defects; the treated colon segment was of normal diameter, and only small adhesions were present. CONCLUSION BPC 157 is a fundamental treatment that quickly restores blood supply to the ischemically injured area and rapidly activates collaterals. This effect involves the NO system.

Membrane of Candida albicans as a target of berberine

BackgroundWe investigated the mechanisms of anti-Candida action of isoquinoline alkaloid berberine, active constituent of medically important plants of Barberry species.MethodsThe effects on membrane, morphological transition, synthesis of ergosterol and the consequent changes in membrane permeability have been studied. Polarization and lipid peroxidation level of the membrane following berberine treatment have been addressed.ResultsMinimal inhibitory concentration (MIC) of berberine against C. albicans was 17.75xa0μg/mL. Cytotoxic effect of berberine was concentration dependent, and in sub-MIC concentrations inhibit morphological transition of C. albicans cells to its filamentous form. Results showed that berberine affects synthesis of membrane ergosterol dose-dependently and induces increased membrane permeability causing loss of intracellular material to the outer space (DNA/protein leakage). Berberine also caused membrane depolarization and lipid peroxidation of membrane constituents indicating its direct effect on the membrane. Moreover, ROS levels were also increased following berberine treatment indicating further the possibility of membrane damage.ConclusionBased on the obtained results it seems that berberine achieves its anti-Candida activity by affecting the cell membrane.

Stable gastric pentadecapeptide BPC 157 and bupivacaine

Bupivacaine toxicity following accidental overdose still lacks therapeutic solution. However, there are major arguments for testing BPC 157 against bupivacaine toxicity in vivo in rats, in particular, and then finally, in vitro. These are: the lack of any known BPC 157 toxicity, a lifesaving effect via the mitigation of arrhythmias in rats underwent hyperkalemia or digitalis toxicity, the elimination of hyperkalemia and arrhythmias in rats underwent succinylcholine toxicity and finally, the reduction of potassium-induced depolarization in vitro (in HEK293 cells) in severe hyperkalemia. Most importantly, BPC 157 successfully prevents and counteracts bupivacaine cardiotoxicity; BPC 157 is effective even against the worst outcomes such as a severely prolonged QRS complex. Here, rats injected with bupivacaine (100mg/kg IP) exhibited bradycardia, AV-block, ventricular ectopies, ventricular tachycardia, T-wave elevation and asystole. All of the fatalities had developed T-wave elevation, high-degree AV-block, respiratory arrest and asystole. These were largely counteracted by BPC 157 administration (50µg/kg, 10µg/kg, 10ng/kg, or 10pg/kg IP) given 30min before or 1min after the bupivacaine injection. When BPC 157 was given 6min after bupivacaine administration, and after the development of prolonged QRS intervals (20ms), the fatal outcome was markedly postponed. Additionally, the effect of bupivacaine on cell membrane depolarization was explored by measuring membrane voltages (Vm) in HEK293 cells. Bupivacaine (1mM) alone caused depolarization of the cells, while in combination with BPC 157 (1µm), the bupivacaine-induced depolarization was inhibited. Together, these findings suggest that the stable gastric pentadecapeptide BPC 157 should be a potential antidote for bupivacaine cardiotoxicity.

Class side effects: decreased pressure in the lower oesophageal and the pyloric sphincters after the administration of dopamine antagonists, neuroleptics, anti-emetics, L-NAME, pentadecapeptide BPC 157 and L-arginine

The ulcerogenic potential of dopamine antagonists and l-NAME in rats provides unresolved issues of anti-emetic neuroleptic application in both patients and experimental studies. Therefore, in a 1-week study, we examined the pressures within the lower oesophageal and the pyloric sphincters in rats [assessed manometrically (cm H2O)] after dopamine neuroleptics/prokinetics, l-NAME, l-arginine and stable gastric pentadecapeptide BPC 157 were administered alone and/or in combination. Medication (/kg) was given once daily intraperitoneally throughout the 7xa0days, with the last dose at 24xa0h before pressure assessment. Given as individual agents to healthy rats, all dopamine antagonists (central [haloperidol (6.25xa0mg, 16xa0mg, 25xa0mg), fluphenazine (5xa0mg), levomepromazine (50xa0mg), chlorpromazine (10xa0mg), quetiapine (10xa0mg), olanzapine (5xa0mg), clozapine (100xa0mg), sulpiride (160xa0mg), metoclopramide (25xa0mg)) and peripheral(domperidone (10xa0mg)], l-NAME (5xa0mg) and l-arginine (100xa0mg) decreased the pressure within both sphincters. As a common effect, this decreased pressure was rescued, dose-dependently, by BPC 157 (10xa0µg, 10xa0ng) (also note that l-arginine and l-NAME given together antagonized each other’s responses). With haloperidol, l-NAME worsened both the lower oesophageal and the pyloric sphincter pressure, while l-arginine ameliorated lower oesophageal sphincter but not pyloric sphincter pressure, and antagonized l-NAME effect. With domperidone, l-arginine originally had no effect, while l-NAME worsened pyloric sphincter pressure. This effect was opposed by l-arginine. All these effects were further reversed towards a stronger beneficial effect, close to normal pressure values, by the addition of BPC 157. In addition, NO level was determined in plasma, sphincters and brain tissue. Thiobarbituric acid reactive substances (TBARS) were also assessed. Haloperidol increased NO levels (in both sphincters, the plasma and brain), consistently producing increased TBARS levels in the plasma, sphincters and brain tissues. These effects were all counteracted by BPC 157 administration. In conclusion, we revealed that BPC 157 counteracts the anti-emetic neuroleptic class side effect of decreased pressure in sphincters and the dopamine/NO-system/BPC 157 relationship.

BPC 157 antagonized the general anaesthetic potency of thiopental and reduced prolongation of anaesthesia induced by L-NAME/thiopental combination.

AimWe hypothesized that certain effects of the general anaesthetic thiopental are dependent on NO-related mechanisms, which were consequently counteracted by stable gastric pentadecapeptide BPC 157.Main methods(1) All rats intraperitoneally received thiopental (20, 30, 40, and 50xa0mg/kg) while medication BPC 157 (10xa0μg/kg, 10xa0ng/kg, and 10xa0pg/kg) was given intraperitoneally at 5xa0min before thiopental. (2) To determine NO-related mechanisms, all rats received intraperitoneally thiopental 40xa0mg/kg while BPC 157 (10xa0μg/kg), l-NAME (10xa0mg/kg) and l-arginine (30xa0mg/kg) were applied alone and/or combined. BPC 157 was given at 25xa0min before thiopental while l-NAME, l-arginine, alone and/or combined, were applied at 20xa0min before thiopental.Key findings(1) BPC 157 own effect on thiopental anaesthesia: BPC 157 (10xa0ng/kg and 10xa0μg/kg) caused a significant antagonism of general anaesthesia produced by thiopental with a parallel shift of the dose–response curve to the right. (2) l-NAME-l-arginine-BPC 157 interrelations: l-NAME: Thiopental-induced anaesthesia duration was tripled. l-arginine: Usual thiopental anaesthesia time was not influenced. Active only when given with l-NAME or BPC 157: potentiating effects of l-NAME were lessened, not abolished; shortening effect of BPC 157: abolished. BPC 157 andl-NAME: Potentiating effects of l-NAME were abolished. BPC 157 andl-NAME andl-arginine: BPC 157xa0+l-NAMExa0+l-arginine rats exhibited values close to those in BPC 157 rats.SignificanceThiopental general anaesthesia is simultaneously manipulated in both ways with NO system activity modulation, l-NAME (prolongation) and BPC 157 (shortening/counteraction) and l-arginine (interference with l-NAME and BPC 157).

Neuroprotective effect of zolpidem against glutamate-induced toxicity is mediated via the PI3K/Akt pathway and inhibited by PK11195

Excitotoxicity is a pathological process in which neuronal dysfunction and death are induced by excessive glutamate stimulation, the major fast excitatory neurotransmitter in the mammalian brain. Excitotoxicity-induced neurodegeneration is a contributing factor in ischemia-induced brain damage, traumatic brain injury, and various neurodegenerative diseases. It is triggered by calcium overload due to prolonged over-activation of ionotropic N-methyl-d-aspartate (NMDA) receptors. Enhanced Ca2+ release results in neuronal vulnerability through several intertwined mechanisms, including activation of proteolytic enzymes, increased production of reactive oxygen species (ROS), mitochondrial dysfunction and modulation of intracellular signalling pathways. We investigated the neuroprotective effect of hypnotic zolpidem, a drug that exerts its central effects at the GABAA receptor complex, against glutamate-induced toxicity in P19 neurons. Zolpidem prevented death of P19 neurons exposed to glutamate, and abolished the glutamate-induced increase in ROS production, p53 and Bax expression, and caspase-3/7 activity. Zolpidem effects were mediated by marked over-activation of Akt kinase. The pro-survival effect, as well as the pAkt induction, were prevented in the presence of wortmannin, an inhibitor of phosphatidylinositol-3-kinase (PI3K) that functions upstream of Akt. The beneficial effect of zolpidem on neuronal survival was not prevented by flumazenil, a GABAA receptor antagonist. PK11195, a drug that modulates the mitochondrial translocator protein 18u202fkDa (TSPO) and F0F1-ATPase, prevented the beneficial effect of zolpidem, indicating that the mechanism of zolpidem action involves preservation of mitochondrial function and integrity. Zolpidem effects were further mediated by prevention of glutamate-induced increase in the expression of the NR2B subunit of NMDA receptor. The obtained results suggest the promising therapeutic potential of zolpidem against excitotoxic insults and highlight the importance of mitochondria and the Akt pathway as valuable targets for therapeutic interventions in glutamate-mediated neuropathological conditions.

The Components Responsible for the Antimicrobial Activity of Propolis from Continental and Mediterranean Regions in Croatia

Propolis is a popular subject of research worldwide due to its therapeutic potential. The antimicrobial activity of propolis appears to be promising but depends on many variables related to its origin such as the content of phenolics and flavonoids. To address this issue with Croatian propolis, which has two major origins Mediterranean and continental), we exposed bacteria (Escherichia coli and Staphylococcus aureus) and yeasts (Candida albicans and Aspergillus niger) to different propolis concentrations (two-fold microdilution method with TCC/formazan endpoint). Total phenolic and flavonoid content and chromatographic profile along with antioxidant activity were assessed. The majority of the 24 propolis samples tested exhibited potent antimicrobial activity against S. aureus bacteria and the yeast C. albicans. Most propolis samples also exhibited robust antioxidative capacity which correlated polyphenol and flavonoid content. To the best of our knowledge this is the first study in which the antimicrobial activity of Croatian propolis is correlated with its constituents.

Probiotics as an adjuvant therapy in major depressive disorder.

Background Major depressive disorder is a common, debilitating psychiatric disorder, which originates from the interaction of susceptibility genes and noxious environmental events, in particular stressful events. It has been shown that dysregulation of hypothalamus-pituitary-adrenal (HPA) axis, imbalance between anti- and pro-inflammatory cytokines, depletion of neurotransmitters (serotonin, norepinephrine and/or dopamine) in the central nervous system, altered glutamatergic and GABAergic transmission have an important role in the pathogenesis of depression. Due to numerous diverse biological events included in the pathophysiology of depression a large number of antidepressant drugs exerting distinct pharmacological effects have been developed. Nevertheless, clinical needs are still not solved. Results Relatively new research strategies advanced the understanding of psychiatric illness and their connections with disturbances in gastrointestinal tract. The existence of bidirectional communication between the brain and the gut has been proven, and an increasing body of evidence supports the hypothesis that cognitive and emotional processes are influenced through the brain-gut axis. On the other hand, microbiome may influence brain function and even behavior giving to the specific microorganisms a psychobiotic potential. Conclusions In this review we discuss the possibilities of classical antidepressant drug treatment being supported with the psychobiotics/probiotic bacteria in patients suffering from major depressive disorder.