Interaction of hypoxia and nicotine acetylcholine receptor signaling network reveals a novel mechanism for lung adenocarcinoma progression in never-smokers

High incident of lung cancer among never smokers and their disease pathogenesis is an unexplained phenomenon. We have analyzed 1727 lung cancer patient data to understand the impact of smoking on overall survival of lung cancer patients and have observed a difference of only 47 days between smokers and never smokers in adenocarcinoma patients suggesting that the disease is equally fatal in never-smokers irrespective of gender. In this study, we have investigated the possible collaboration between the nAChR and hypoxia signaling pathway to elucidate a mechanism of disease progression in never-smokers. We report a previously unidentified increase in both acetylcholine and nAChR-α7 levels in non small cell lung cancer cells in hypoxia. Similar increase in ubiquitously expressed nAChR-α7 transcripts was also observed in other cancer lines. A direct binding of HIF-1α with the hypoxia response element (HRE) present at -48 position preceding the transcriptional start site in nAChR-α7 promoter region was established. Significantly, the increased acetylcholine levels in hypoxia drove a feedback loop via modulation of PI3K/AKT pathway to stabilize HIF-1α in hypoxia. Further, Bungarotoxin, an antagonist of nAChR-α7 significantly reversed hypoxia mediated metastasis and induction of HIF-1α in these cells. Our study gives a plausible explanation for the equally worse prognosis of lung adenocarcinoma in never-smokers wherein the nAChR signaling is enhanced in hypoxia by acetylcholine, in the absence of nicotine.

The Case of a Glucocorticoid-Induced Myopathy & Myasthenia Gravis Combination: Is There a Border?

Introduction: Glucocorticoid-induced myopathy is the sign of Cushing’s syndrome in 83%, but can also be consequence of exogenous administration of glucocorticoids. The patients with the compromised neuromuscular system (for example, because of myasthenia gravis) have a higher risk of this complication. Differential diagnosis of glucocorticoid-induced myopathy is challenging. Clinical case. A 77-year woman complained of difficulty in the act of breathing, swallowing, weakness in the limbs, inability to self-service, ptosis. Due to the growing complaints the patient was hospitalized to ICU. From anamnesis: She was diagnosed with myasthenia gravis 2 months ago. The presence of autoantibodies to the nicotine acetylcholine receptor was confirmed. Pyridostigmine bromide 60 mg/day, Methylprednisolone 20 mg/day were prescribed for 5 days. The patient noted some improvement. After the next neurologist’s examination the dose of Methylprednisolone was doubled. (It is not known for certain whether the doctor wanted to reduce the dose and the patient misunderstood.) She noted some worsening: weakness and speech difficulties increased. She returned to the previous dose after 3 days, but there was no improvement. Objectively: hypersthenic body type, BMI 39 kg/m2, predominant deposition of adipose tissue in the abdomen and face. Breathing without a ventilator. BP 250/100 mm Hg, heart rate 87 BPM. Laboratory tests revealed hyperkalemia of 8.76 mmol/l (3.5–5), creatinine of 481 mmol/l (44–124), hyperglycemia of 16 mmol/l. Glucocorticoid-induced myopathy was suspected, the administration of methylprednisolone was cancelled. The improvement of the condition was noted in 3 days. Blood pressure, glycaemia, levels of potassium and creatinine returned to normal. A biopsy of muscle revealed: atrophy of type I and II muscle fibers. There were no signs of necrosis, regeneration or inflammation. Thereby the diagnosis of glucocorticoid-induced myopathy was confirmed. Conclusion. Glucocorticoid-induced myopathy may look like a decompensation of neuromuscular disease. Since there are no accurate diagnostic tests nowadays the main argument of diagnosis is the paradoxical reaction after increasing the dose of glucocorticoids as well in this case. Even a short period of use of these drugs can lead to the development of side effects. It is necessary to regularly monitor the dynamics of the condition of such patients.

The Study on Neural Remodeling of Medullary Visceral Zone in Early Sepsis and Interfered by Cholinergic Anti-Inflammatory Pathway ​

BackgroundStudies including our own have shown that the Medullary Visceral Zone (MVZ) can effectively regulate systemic inflammation and immunity through the Cholinergic Anti-inflammatory Pathway (CAP). Sepsis usually causes neuroinflammation in the Central Nervous System (CNS), which will inevitably affect the structure and function in related brain areas such as MVZ, whether the intervention of CAP can affect the structure and function of the MVZ in sepsis needs to be further verified. Methods64 adults, specific pathogens free Sprague-Dawley male rats were used in this study. The septic models were prepared by cecum ligation and puncture (CLP) method, GTS-21 (a selective α7 nicotinic acetylcholine receptor agonist which can mimic CAP’s activation) and MLA (a powerful and selective nicotine acetylcholine receptor antagonist which can mimic CAP’s blocking) were used to interfere CAP. The pathological changes, apoptosis, the expressions of Tyrosine Hydroxylase (TH) and Choline acetyltransferase (CHAT), the expression levels of GAP-43 mRNA, Olig-2 mRNA, VEGF mRNA, GFAP mRNA, MMP-9 mRNA in MVZ were analyzed among different groups. ResultsIn this study, we found that sepsis induced apoptosis and functional suppression of catecholaminergic and cholinergic neurons and gliosis in MVZ, up-regulated key genes’ expressions such as GAP-43 mRNA，GFAP mRNA，VEGF mRNA，MMP-9 mRNA, down-regulated the expression of Olig-2 mRNA. GTS-21, a selective α7 nicotinic acetylcholine receptor agonist, obviously mitigated the above changes; whereas, methyllycaconitine (MLA), a powerful and selective nicotine acetylcholine receptor antagonist, significantly aggravated these changes. ConclusionsOur research shows that activating CAP can effectively mitigate the neural remodeling and neuronal suppression induced by early sepsis in MVZ, the mechanism may involve with its control of systemic and local inflammation. This study reveals that MVZ and CAP may be potential targets to curb the inflammatory storm in early sepsis.

PROINSECTICIDES

Pro-insecticides are chemical compounds, the bioactivation of which occurs in the body of insects, where they are transformed into metabolites with greater insecticidal activity. These substances account for 20% of the total amount of insecticides on the market, and they account for 34% of the value of all world sales. Only after many years of use of insecticides did it become known that a significant part of them are precursors. According to the Insecticide Resistance Action Committee (IRAC), about 40% of chemical groups contain precursors that require structural changes to manifest their insecticidal properties. Currently, 16 chemical groups of insecticides are known, in which there are representatives of pro-insecticides. The main molecular targets are: nicotine-acetylcholine receptor (NAChR), voltage-gated K- and Na-channels, Cl-channel of the GABAreceptor, Cl-channel of glutamate receptor, acetylcholinesterase (AChE) and ryanodine receptor (RyR). The main direction of using pro-insecticides is the control of insecticide-resistant insect populations. The review summarizes and analyzes modern data on pro-insecticides, describes the main representatives and the mechanism of their transformation in the insect organism. Key words: pro-insecticides, insecticide resistance, insecticide mode of action, bioactivation

Molecular Regulation of α3β4 Nicotinic Acetylcholine Receptors by Lupeol in Cardiovascular System

Cardiovascular disease (CVD) occurs globally and has a high mortality rate. The highest risk factor for developing CVD is high blood pressure. Currently, natural products are emerging for the treatment of hypertension to avoid the side effects of drugs. Among existing natural products, lupeol is known to be effective against hypertension in animal experiments. However, there exists no study regarding the molecular physiological evidence against the effects of lupeol. Consequently, we investigated the interaction of lupeol with α3β4 nicotinic acetylcholine receptors (nAChRs). In this study, we performed a two-electrode voltage-clamp technique to investigate the effect of lupeol on the α3β4 nicotine acetylcholine receptor using the oocytes of Xenopus laevis. Coapplication of acetylcholine and lupeol inhibited the activity of α3β4 nAChRs in a concentration-dependent, voltage-independent, and reversible manner. We also conducted a mutational experiment to investigate the influence of residues of the α3 and β4 subunits on lupeol binding with nAChRs. Double mutants of α3β4 (I37A/N132A), nAChRs significantly attenuated the inhibitory effects of lupeol compared to wild-type α3β4 nAChRs. A characteristic of α3β4 nAChRs is their effect on transmission in the cardiac sympathetic ganglion. Overall, it is hypothesized that lupeol lowers hypertension by mediating its effects on α3β4 nAChRs. The interaction between lupeol and α3β4 nAChRs provides evidence against its effect on hypertension at the molecular-cell level. In conclusion, the inhibitory effect of lupeol is proposed as a novel therapeutic approach involving the antihypertensive targeting of α3β4 nAChRs. Furthermore, it is proposed that the molecular basis of the interaction between lupeol and α3β4 nAChRs would be helpful in cardiac-pharmacology research and therapeutics.

JN403, an alpha-7-nicotine-acetylcholine-receptor agonist, reduces alpha-synuclein induced inflammatory parameters of in vitro microglia but fails to attenuate the reduction of TH positive nigral neurons in a focal alpha-synuclein overexpression mouse model of Parkinson’s disease

Alpha-7-nicotine-acetylcholine-receptor (α7-nAChRs) agonists modulate the cholinergic antiinflammatory pathway to attenuate proinflammatory signals and reduce dopaminergic neuronal cell loss in toxin-induced experimental murine models of Parkinson’s disease (PD). The protein α-synuclein (αSyn) is considered to represent the major pathogenic component in the etiology and progression of sporadic PD. However, no research has been performed to evaluate the effect of α7-nAChR agonists in human αSyn mediated models of PD. We, therefore, investigated the effect of the compound JN403, an α7-nAChR specific agonist, in αSyn treated in vitro microglia culture and in a human αSyn overexpression in vivo mouse model. In primary mouse microglia cells, αSyn fragment 61-140 treatment increased the release of nitric oxide (NO), tumor necrosis factor (TNF)-α and interleukin (IL)-6, and decreased cell viability. In contrast, 100 nM or 1 μM of JN403 co-incubation significantly reduced the level of NO and TNF-α release in the microglial cells. For in-vivo testing of JN403, a recombinant adeno-associated viral vector (rAAV)-mediated unilateral intranigral overexpression of human wild-type-αSyn (WT-αSyn) or of the control protein luciferase (luc) was induced via stereotactic delivery in C57/BL6N mice. Targeted WT-αSyn overexpression reduced 20% of the number of tyrosine hydroxylase (TH) positive (+) nigral neurons after 10 weeks. Subcutaneous daily treatment of 30 mg/kg JN403 over 9 weeks starting at postoperative week 1 did not alter the decrease of TH+ neuronal numbers, and microglial density in WT-αSyn overexpression mouse model. The reduced density of TH+ striatal terminals in the WT-αSyn groups was also not recovered by the JN403 treatment. In summary, JN403, an α7-nAChR specific agonist shows a beneficial effect on ameliorating proinflammatory signals in αSyn exposed microglia cells. However, no significant in-vivo treatment effect was found in an intranigral WT-αSyn overexpression mouse model of PD.