Role In Development Of Tolerance Research Articles

Influence of forkhead box protein 3 gene polymorphisms in recurrent pregnancy loss: A meta-analysis

BackgroundTreg cells play an important role in development of tolerance in maternal immune system against the semi-allogenic embryo. Human forkhead box protein 3 (FOXP3) gene, is the major transcription factor responsible for the regulation of Treg function during pregnancy. Single nucleotide polymorphisms (SNPs) of FOXP3 gene have been reported as a risk factor for Recurrent Pregnancy Loss (RPL), however, results from previous studies are inconsistent. MethodologyWe have collected data from different studies to investigate the overall association of FOXP3 SNPs with risk of RPL. PubMed, Google Scholar, Elsevier, and Cochrane databases were searched to identify eligible studies. Odds Ratio (OR) and 95 % Confidence Interval (CI), calculated via fixed effect or random effect models, were used to evaluate strength of association. This meta-analysis included 11 studies (1383 RPL cases and 1413 controls) of 6 SNPs: rs3761548 A/C, rs2232365 A/G, rs2294021 T/C, 2280883 T/C, rs5902434del/ATT and rs141704699C/T, with ≥2 studies per SNPs and at least 1 significant result. ResultsWe observed that FOXP3 polymorphism was predominantly present in Asian women with history of RPL. rs2232365 A/G, rs3761548 A/C, rs2294021 T/C, rs2280883 T/C and rs5902434del/ATT polymorphisms were significantly associated with risk of RPL in Indian population. Further, among the most commonly seen polymorphism, rs3761548 A/C was significantly associated with risk of RPL in women from Kazakhstan, China and Gaza, Palestine; rs2232365 A/G in populations of Kazakhstan, Egypt, Iran and Gaza, Palestine. Results of this study indicates that FOXP3 polymorphism is significantly associated with risk of RPL, especially in Asians.

Effect of Forkhead Box Protein 3 Gene Polymorphisms in Recurrent Pregnancy Loss: A meta-analysis

Treg cells play an important role in development of tolerance in maternal immune system against the semi-allogenic embryo. Human forkhead box protein 3 (FOXP3) gene, is the major transcription factor responsible for the regulation of Treg function during pregnancy. Single nucleotide polymorphisms (SNPs) of FOXP3 gene have been reported as a risk factor for Recurrent Pregnancy Loss (RPL), however, results from previous studies are inconsistent. In this meta-analysis, we collected data from different studies to investigate the overall association of FOXP3 SNPs with risk of RPL. PubMed, Google Scholar, Elsevier, and Cochrane databases were searched to identify eligible studies. Odds Ratio (OR) and 95% Confidence Interval (CI), calculated via fixed effect or random effect models, were used to evaluate strength of association. This meta-analysis included 11 studies (1383 RPL cases and 1413 controls) of 6 SNPs: rs3761548 A/C, rs2232365 A/G, rs2294021 T/C, 2280883 T/C, rs5902434del/ATT and rs141704699 C/T, with ≥ 2 studies per SNPs and at least 1 significant result. We observed that FOXP3 polymorphism was predominantly present in Asian women with history of RPL. rs2232365 A/G, rs3761548 A/C, rs2294021 T/C, rs2280883 T/C and rs5902434del/ATT polymorphisms were significantly associated with risk of RPL in Indian population. Further, among the most commonly seen polymorphism, rs3761548 A/C was significantly associated with risk of RPL in women from Kazakhstan, China and Gaza, Palestine; rs2232365 A/G in populations of Kazakhstan, Egypt, Iran and Gaza, Palestine. Results of this study indicates that FOXP3 polymorphism is significantly associated with risk of RPL, especially in Asians.

Membrane Transporters of the Major Facilitator Superfamily Are Essential for Long-Term Maintenance of Phenotypic Tolerance to Multiple Antibiotics in E. coli.

ABSTRACTAntibiotic tolerance is not only the key underlying the cause of recurrent and chronic bacterial infections but it is also a factor linked to exacerbation of diseases, such as tuberculosis, cystic fibrosis-associated lung infection, and candidiasis. This phenomenon was previously attributed to a switch to physiological dormancy in a bacterial subpopulation triggered by environmental signals. However, we recently showed that expression of phenotypic antibiotic tolerance during nutrient starvation is highly dependent on robust production of proteins that actively maintain the bacterial transmembrane proton motive force (PMF), even under a nutrient-deficient environment. To investigate why PMF needs to be maintained for expression of phenotypic antibiotic tolerance, we tested the relative functional role of known transporters and efflux pumps in tolerance development by assessing the effect of deletion of specific efflux pump and transporter-encoding genes on long-term maintenance of antibiotic tolerance in an Escherichia coli population under starvation. We identified eight specific efflux pumps and transporters and two known efflux pump components, namely, ChaA, EmrK, EmrY, SsuA, NhaA, GadC, YdjK, YphD, TolC, and ChaB, that play a key role in tolerance development and maintenance. In particular, deletion of each of the nhaA and chaB genes is sufficient to totally abolish the tolerance phenotypes during prolonged antimicrobial treatment. These findings therefore depict active, efflux-mediated bacterial tolerance mechanisms and facilitate design of intervention strategies to prevent and treat chronic and recurrent infections due to persistence of antibiotic-tolerant subpopulations in the human body.IMPORTANCE We recently showed that the antibiotic-tolerant subpopulation of bacteria or persisters actively maintain the transmembrane proton motive force (PMF) to survive starvation stress for a prolonged period. This work further shows that the reason why antibiotic persisters need to maintain PMF is that PMF is required to support a range of efflux or transportation functions. Intriguingly, we found that tolerance-maintaining efflux activities were mainly encoded by 10 efflux or transporter genes. Because our study showed that deletion of even one of these genes could cause a significant reduction in tolerance level, we conclude that the products of these genes play an essential role in enhancing the survival fitness of bacteria during starvation or under other adverse environmental conditions. These gene products are therefore excellent targets for future design of antimicrobial agents that eradicate antibiotic tolerant persisters and prevent occurrence of chronic and recurrent human infections.

Seed priming with endophytic Bacillus subtilis strain-specifically improves growth of Phaseolus vulgaris plants under normal and salinity conditions and exerts anti-stress effect through induced lignin deposition in roots and decreased oxidative and osmotic damages

Bacillus subtilis is one of the non-pathogenic beneficial bacteria that promote plant growth and stress tolerance. In the present study, we revealed that seed priming with endophytic B. subtilis (strains 10–4, 26D) improved Phaseolus vulgaris L. (common bean) seed germination and plant growth under both saline and non–saline conditions. 10–4 and 26D decreased oxidative and osmotic damage to the plant cells since bacterial inoculations reduced lipid peroxidation and proline accumulation in plants under salinity. 26D and especially 10–4 preserved different elevated levels of chlorophyll (Chl) a and Chl b in bean leaves under salinity, while carotenoids (Car) increased only by 10–4 and slightly decreased by 26D. Under normal conditions, 10–4 and 26D did not affect Chl a and Car concentrations, while Chl b decreased in the same plants. Under non–saline and especially saline conditions, 10–4 and 26D significantly increased lignin accumulation in plant roots and the highest lignin content along with better growth and oxidative damages reduction was observed after 10–4 inoculation under salinity, indicating a major role of B. subtilis–induced strengthening the root cell walls in the implementation protective effect of studied bacteria on plants. Therefore, B. subtilis 10–4 and 26D exerts protective effects on the growth of common bean plants under salinity by regulating plant defense mechanisms and the major role in tolerance development may contribute through the activation by B. subtilis lignin deposition in roots. The obtained data also indicates a strain-dependent efficiency of endophytic B. subtilis since strains 10–4 and 26D differently improved growth attributes and modulates cellular response reactions of the same common bean plants both under normal and salinity conditions, that generates interest for further investigations in this direction.

Assessing Opioid Tolerance Mechanisms in an Isolated Murine Dorsal Root Ganglia Neuron Model

Our current understanding of tolerance development to opioids has largely been collected through whole‐animal studies and cell culture models, yet many questions related to the underlying mechanisms remain. One approach our lab has employed is the use of isolated dorsal root ganglia (DRG) neurons from adult mice as an ex vivo native cell culture model to study the effects of opioids on neurons and to further our understanding of tolerance development within a single cell. Our initial studies focused on comparing morphine and oxycodone, two opioids widely used clinically for the treatment of pain. Interestingly, oxycodone and morphine are known to differ in antinociceptive potency and oral bioavailability when tested in whole animals, yet we found using whole‐cell patch clamp electrophysiology that acute oxycodone and morphine had similar potencies in DRG neurons. A significant decrease in neuronal excitability as measured by an increase in the potential required to fire an action potential (i.e. threshold potential) from baseline was observed following the application of 3 μM oxycodone (p < 0.01) or morphine (p < 0.05). Marked increases in threshold potential occurred rapidly, within five to ten minutes of drug application. When incubated in media containing 3 μM morphine, DRG neurons displayed tolerance development to a morphine challenge two days later as indicated by no change in threshold potential (p < 0.05). Tolerance was observed following overnight incubation in media containing 10 μM of either morphine or oxycodone (p < 0.05). β‐arrestin 2 knockout (β‐arr2 KO) mice do not develop antinociceptive tolerance to morphine in vivo, and we found that DRG neurons isolated from β‐arr2 KO mice and incubated in 10 μM morphine continued to respond to a 3 μM morphine challenge, which showed that tolerance to morphine did not develop in a single‐cell preparation (p < 0.05). We further observed that overnight oxycodone incubation did not produce tolerance in β‐arr2 KO DRGs (p < 0.05), providing evidence that β‐arrestin 2 plays a role in tolerance development to both oxycodone and morphine. Our lab has also shown a substantial role of protein kinase C (PKC) in morphine antinociceptive tolerance in vivo using warm water tail withdrawal studies. We tested an inhibitor of PKC, Bisindolylmaleimide XI, on DRG neurons incubated overnight with 10 μM oxycodone and found a rapid reversal of tolerance to oxycodone (p < 0.01). These data indicate that PKC‐mediated mechanisms also underlie oxycodone tolerance, similarly to that of morphine tolerance, and operate at the single‐cell level. Opioid tolerance mechanisms are undoubtedly complicated and involve multiple signaling molecules, including β‐arrestin 2 and PKC. DRG neurons are a solid model bridging in vivo and in vitro studies to investigate cellular mechanisms of opioid tolerance.Support or Funding Information5T32DA007027P30DA033934This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Effect of chronic ethanol exposure and its withdrawal on the endocannabinoid system

The present study investigated the effect of ethanol (EtOH) exposure and its withdrawal on the central endocannabinoid system utilizing an EtOH vapor inhalation model, which is known to produce functional tolerance and dependence to EtOH. Swiss Webster mice ( n = 24) were exposed to EtOH vapors for 72 h. Mice were sacrificed after 72 h following EtOH exposure ( n = 12) and 24 h after its withdrawal ( n = 12). Radioligand binding assays were performed to measure the density of CB 1 receptor and CB 1 receptor agonist-stimulated [ 35S]GTPγS binding in crude synaptic membranes isolated from the cortex, hippocampus, striatum and cerebellum. The density of CB 1 receptor was significantly decreased (31–39%) in all the brain regions when compared to the control group. The CB 1 receptor-stimulated G i/o protein activation was also found to be decreased (29–40%) in these brain regions of EtOH exposed mice. Recovery of the CB 1 receptor density, in addition to, the CB 1 receptor-mediated G-protein activation was observed after 24 h withdrawal from EtOH. The levels of cortical anandamide, which was significantly increased (147%) by EtOH exposure, returned to basal levels after 24 h of withdrawal from EtOH exposure. A significant reduction (21%) in the activity of fatty acid amide hydrolase was found in the cortex of EtOH administered mice. Taken together, the neuroadaptation in the EC system may have a potential role in development of tolerance and dependence to EtOH.

Roles of Superoxide, Peroxynitrite, and Protein Kinase C in the Development of Tolerance to Nitroglycerin

A current hypothesis states that tolerance to nitroglycerin (GTN) involves increased formation of superoxide (O2*-). Studies showing that inhibitors of protein kinase C (PKC) prevent tolerance to GTN suggest the involvement of PKC activation, which can also increase O2*-. We examined the roles of O2*-, peroxynitrite (ONOO-), and PKC activation in GTN tolerance. Pre-exposure of rat aortic rings to GTN (5 x 10(-4) M) for 2 h caused tolerance to the vasodilating effect of GTN, as evidenced by a substantial rightward shift of GTN concentration-relaxation curves. This shift was reduced by treatment of the rings with the antioxidants uric acid, vitamin C, or tempol or the PKC inhibitor chelerythrine. We also found that O2*- generation via xanthine/xanthine oxidase in the bath induced tolerance to GTN. However, responses to nitroprusside were not affected. In vivo tolerance produced in rats by 3-day i.v. infusion of GTN was also almost completely prevented by coinfusion of tempol. In bovine aortic endothelial cells (EC), addition of GTN produced a marked increase in tyrosine nitrosylation, indicating increased ONOO- formation. This action was blocked by prior treatment with uric acid, superoxide dismutase, NG-nitro-L-arginine methyl ester, or chelerythrine. We also demonstrated that GTN translocates the alpha- and epsilonPKC isoforms in EC. However, PKCzeta was not affected by GTN treatment. In conclusion, tolerance to GTN involves enhanced production of O2*- and ONOO- and activation of NO synthase. Furthermore, sustained activation of alpha- and epsilonPKC isozymes in EC by GTN may play a role in development of tolerance.

Tolerance to nitroglycerin in rabbit aorta: Investigating the involvement of the mu isozyme of glutathione S-transferases

We have proposed that glutathione S-transferases (GSTs), especially the mu isozyme, play a critical role in the metabolism of nitroglycerin (glyceryl trinitrate, GTN), leading to pharmacologic effects. Here we study this enzyme(s) during tolerance development in male New Zealand white rabbits. Each aorta was divided into two segments designated as GTN pretreated and buffer control. Tolerance was induced in rabbit aortic strips so assigned by incubation with GTN (0.22 mM). The activity of the mu isozyme and of total GSTs was determined in portions of each segment. In each rabbit aorta, the response to GTN (0.5 μM) was determined in GTN-pretreated and buffer-pretreated strips by measuring cyclic GMP levels (N = 7 pairs) and percent relaxation (N = 4 pairs). In GTN-pretreated strips, a significant decrease was observed in the activity of the mu isozyme of GST, while the total GST activity was unchanged as compared with control strips. The decrease in isozyme activity correlated very well with the decrease in response to GTN. Two rabbit aortae did not become tolerant, and the activity of the mu isozyme was also not affected. The levels of thiols were not affected by GTN pretreatment and aortae tolerant to GTN did not develop tolerance to S-nitroso acetylpenicillamine (SNAP), indicating that thiol depletion and guanylate cyclase desensitization probably play a minor role in tolerance development to GTN in our model. These studies suggest that tolerance to GTN in rabbit aorta in vitro is associated with a decrease in GST mu activity, which correlates well with the decrease in GTN response.

The NMDA receptor antagonist MK-801 prevents long-lasting non-associative morphine tolerance in the rat

Several studies have demonstrated that the N-methyl-D-aspartate (NMDA) antagonist MK-801 attenuates the development of morphine tolerance and withdrawal. These studies employed repeated morphine injections to induce tolerance, a procedure in which learning has been suggested to play a significant role in tolerance development. MK-801 has been reported to block some types of learning, and it is unclear, therefore, whether the effect of MK-801 on tolerance development is due to its antagonism of associative (learning) or non-associative factors. Moreover, previous studies have tested the effects of MK-801 on morphine tolerance only up to 48 h after its induction; yet morphine tolerance can persist for many months, and it is not known whether MK-801 can block long-lasting tolerance. In the present study, therefore, we adopted a model of morphine tolerance in which the involvement of learning is minimized by using a single injection of morphine in a sustained-release preparation, and we tested tolerance for up to 56 days. In the first experiment, simultaneously administering MK-801 (0.2 mg/kg, s.c.) and morphine (60 mg/kg, s.c.), each in a sustained-release preparation, abolished tolerance that lasted at least 12 days. Analgesia was measured in the hot-plate test following a test dose of morphine (15 mg/kg, i.p.). In the second experiment, delivering MK-801 and morphine as before, the duration of morphine-induced catalepsy and analgesia was prolonged. Nevertheless, 24 h later one symptom of naloxone-precipitated withdrawal was significantly attenuated in these same animals.(ABSTRACT TRUNCATED AT 250 WORDS)

Regional GABA/benzodiazepine receptor/chloride channel coupling after acute and chronic benzodiazepine treatment

GABA/benzodiazepine coupling was evaluated in 8 regions of rat brain by the ability of GABA to stimulate 0.5 nM [ 3H]flunitrazepam binding. Rats were treated acutely with diazepam (p.o) or chronically with flurazepam, offered in the drinking water for 4 weeks, and compared to a pair-handled vehicle-treated control group. Regional variations in GABA/benzodiazepine coupling were found in control membranes. GABA increased benzodiazepine binding maximally (40%) in cerebellum and medulla, and least (25%) in olfactory bulb. A significant decrease in the effect of GABA was found in cortex of chronically treated rats immediately after, but not 2 days following treatment. The E max for GABA stimulation of [ 3H]flunitrazepam binding was significantly increased in medulla after acute treatment but was not altered after acute or chronic treatment in other brain areas evaluated. Treatment had no effect on the ability of bicuculline to inhibit [ 3H]flunitrazepam binding in cortex. Benzodiazepine /Cl − couping in cortex or hippocampus of acutely and chronically treated rats, evaluated by the ability of Cl − to stimulate specific [ 3H]flunitrazepam binding, was not changed. The results support the hypothesis that a functional uncoupling of the benzodiazepine recognition site from the GABA receptor in cortex, but not from the anion recognition site, may play a role in tolerance development.

Spinal antagonism of tolerance and dependence induced by systemically administered morphine

We observed that the spinal cord of rats was involved in development of tolerance and dependence to morphine that was administered systemically by s.c. morphine-pellet implantation. Rats, surgically fitted with intrathecal catheters, were injected intrathecally (i.t.) with saline or 2.4 nmol β-chlornaltrexamine (β-CNA), an irreversible opiate antagonist. Twenty-four h later, animals were implanted s.c. with either placebo or morphine pellets. Seventy-two h after implanting pellets, development of tolerance or dependence was assessed. Control animals implanted with morphine pellets became tolerant to analgesia induced by i.p. injections of morphine as determined by the use of tail click and hot plate analgesic assays. β-CNA pretreatment antagonized the effects of i.p. injections of morphine and blocked development of tolerance in morphine-implanted animals. Dependence was assessed by observing several characteristic signs of precipitated withdrawal. Treatment with β-CNA before morphine treatment antagonized naloxone-induced expression of withdrawal for all signs observed, except weight loss. We conclude that the spinal cord plays a significant role in development of tolerance and dependence induced by systemically administered opiates.

Tolerance to the antinociceptive effect of morphine in the spinal rat

Tolerance and withdrawal were studied in spinal and intact rats receiving morphine hydrochloride (10–20 mg/kg, i.p.) every 12 hr for 10 days. All rats developed tolerance to morphine-induced antinociception measured by the tail-flick method. Tolerance was not reversed by d,l-5-hydroxytryptophan (200 mg/kg, i.p.). Naloxone (1–2 mg/kg) induced signs of withdrawal both in spinal and intact rats. Thus, functional changes in the aminergic systems and in brain and brainstem structures appear not to be necessary for tolerance to morphine-induced antinociception and for dependence on morphine. The experiment does not support the hypothesis that conditioning to environmental stimuli and changes in cognitive functions play a major role in tolerance development in the rat.

Involvement of 5-hydroxytryptamine in tolerance development in the morphine-induced Straub tail reaction.

The tolerance development in Straub tail reaction [STR] on successive treatments with morphine was studied. Morphine at 20 mg/kg, four times a day, produced tolerance in STR more rapidly than 10 mg/kg, four times a day. Significant tolerance in STR developed when morphine was injected daily into the cerebral ventricle in mice. STR in tolerant mice was markedly reduced by treatment with L-5-hydroxytryptophan compared to that in non-tolerant mice. The cortical 5-hydroxytryptamine [5-HT] content in tolerant mice was significantly decreased compared to that in non-tolerant mice. These results suggest that cortical 5-HT may play an important role in tolerance development in STR.

Effect of p-chlorophenylalanine on the acquisition of tolerance to the hypothermic effects of ethanol.

Daily administration of ethanol in a liquid diet or by intubation were both effective in producing tolerance to the hypothermic effects of ethanol. When p-chlorophenylalanine (p-CPA) was administered in a dose previously demonstrated to maintain extensive brain serotonin (5-HT) depletion, the temperature-lowering effects of ethanol were less pronounced than in corresponding controls. The analysis of the effect of p-CPA on tolerance development took into account both initial body temperature and the degree of hypothermia. This study extends our findings with respect to the inhibitory effect of p-CPA on the development of tolerance to the motor-impairing effects of ethanol, and suggests that 5-HT may have a role in tolerance development to ethanol.

Effect of p-chlorophenylalanine on the acquisition of tolerance to ethanol and pentobarbital.

Rats were rendered tolerant to ethanol or pentobarbital by daily oral administration. Motor impairments after test doses of ethanol or pentobarbital were measured prior to and at various times during chronic treatment in order to assess the degree of tolerance development. Chronic administration of p-chlorophenylalanine (p-CPA) in a dosage regimen which produced and maintained approximately 95% depletion of brain serotonin (5-HT) did not alter motor impairment after initial acute administration of ethanol or pentobarbital. However, the rate of tolerance development to the motor-impairing effects of both drugs was slowed down in p-CPA-treated rats, p-CPA did not appear to exert this effect by altering the disposition of ethanol or pentobarbital, since blood levels determined 20 min after administration of the test doses were similar in animals treated with p-CPA and in controls. These findings suggest that brain 5-HT may have a role in tolerance development to ethanol and pentobarbital.