Activating α7nAChR suppresses systemic inflammation by mitigating neuroinflammation of the medullary visceral zone in sepsis in a rat model.

Our previous studies have shown that activating α7nAChRs suppresses systemic inflammation and immunity through the cholinergic anti-inflammatory pathway (CAP) in early sepsis. Now that the medullary visceral zone (MVZ) is the center of CAP and responsible for regulating systemic inflammation, what changes will occur in MVZ's pathology and function in sepsis, especially when interfering with α7nAChRs? Does activation of MVZ's α7nAChRs contribute to the inhibition of systemic inflammation? To clarify these issues, we explored the systemic inflammation and immunity state by detecting serum levels of TNF-α, IL-6, HMGB1, sCD14, and CD4+CD25+Treg and TH17 lymphocytes percentage, meanwhile, we analyzed the apoptosis of cholinergic and catecholaminergic neurons and the expressions of tyrosine hydroxylase (TH) and choline acetyltransferase (CHAT) in MVZ in sepsis and the interfering effects on α7nAChRs. In this study, we found that in sepsis, serum TNF-α, IL-6, HMGB1, sCD14, CD4+CD25+Treg, and TH17 lymphocytes significantly increased and the ratio of Treg/TH17 significantly decreased, cholinergic and catecholaminergic neurons underwent apoptosis with low expressions of TH and CHAT in MVZ; activation of α7nAChRs not only significantly decreased the levels of septic serum TNF-α, IL-6, HMGB1, sCD14, and TH17 lymphocytes (P ＜ 0.05), but also significantly reduced cholinergic and catecholaminergic neurons' apoptosis, and promoted expressions of TH/CHAT. Our study reveals that sepsis undermines MVZ through neuroinflammation which contributes to the uncontrolled systemic inflammation. Activating central α7nAChRs is not only helpful to restore MVZ's structure and function but also beneficial to subside the inflammatory storm in sepsis. Even if MVZ is damaged in sepsis, cholinergic neurons in MVZ still regulate the systemic inflammation stably.

Neuroinflammation in the medullary visceral zone exert a powerful impaction on the systemic inflammation in sepsis through cholinergic anti-inflammatory pathway.

To investigate whether sepsis-induced neuroinflammation of medulla visceral zone (MVZ) predominates the systemic inflammation through cholinergic anti-inflammatory pathway (CAP), and to explore the effect of central anti-inflammation on systemic inflammation. 112 adult Sprague-Dawley male rats were randomly divided into sepsis experimental group (n = 56) and neuroinflammation experimental group (n = 56). The two experimental groups were individually randomly divided into control group (n = 8), model group (n = 16), central anti-inflammatory group (n = 16) and vagus transection group (n = 16). Rats in two control groups were administered with saline at the dose of 6 mL/kg intraperitoneally or with 25 µL artificial cerebrospinal fluid injected into forth ventricle once a day for 3 days. Rats in two model groups were administered with Lipopolysaccharide (LPS) at the dose of 6 mg/kg intraperitoneally or with 25 µg/25 µL LPS injected into forth ventricle once a day for 3 days. Rats in two central anti-inflammatory groups were fed with 10 mg/mL minocycline sucrose solution as the only water source for 4 days prior to be treated as the model groups of their own, and feeding style was continued until the end of the experiment. Rats in the two vagus transection groups were undergone right vagotomy and 7 days of adaptive feeding prior to be treated as the same as those in the central anti-inflammatory group of their own. The Murine Sepsis Score (MSS), mortality rate and heat rate variability (HRV) were recorded during the last 3 days of intervention. Then the rats were sacrificed and blood samples were collected for ELISA analysis to detect the serum level of inflammatory cytokines such as TNF-α, IL-6, and IL-10. The expression of TNF-α and IL-6 in medulla oblongata were analyzed by Western blot. The correlation and regression analysis among the expression levels of cytokines in medulla oblongata, HRV indexes and serum inflammatory cytokines were performed. The mortality rate and MSS of the sepsis model group and the MVZ's neuroinflammation model group were significantly higher than those of their own control group, and the central anti-inflammation reduced the mortality rate and MSS scores of the two model groups, while the right vagotomy abolished the effect of central anti-inflammatory. In the sepsis model group and the MVZ's neuroinflammation model group, the levels of TNF-α, IL-6, and other cytokines in serum and MVZ were significantly increased, and HRV indexes (SDNN, RMSSD, LF, HF, LF/HF) were significantly decreased (P = 0.000). Central anti-inflammatory treatment reversed the above changes. However, right vagotomy abolished the central anti-inflammatory effect. Correlation and regression analysis showed that there was a significant linear correlation among the expression of inflammatory factors in MVZ, the indexes of HRV and the levels of serum cytokines. Our study shows that sepsis-induced MVZ's neuroinflammation exert a powerful influence on the systemic inflammation through CAP in sepsis. Central anti-inflammation effectively improves systemic inflammation through inhibiting MVZ's neuroinflammation in sepsis. The time domain and frequency domain indexes of HRV can reflect the regulatory effect of CAP and the degree of inflammation of MVZ, which may be potentially used to monitor the condition and treatment effectiveness of sepsis patients.

Sepsis-induced inflammatory demyelination in medullary visceral zone and cholinergic anti-inflammatory pathway: Insights from a Rat's model study.

Background: Our previous studies have demonstrated that the activated Cholinergic Anti-inflammatory Pathway (CAP) effectively suppresses systemic inflammation and immunity in early sepsis. Some parameters of Heart Rate Variability (HRV) could be used to reflect the regulatory activity of CAP. However, in the early stages of severe sepsis of some patients, the inflammatory storm can still result in multiple organs dysfunction and even death, suggesting they lose CAP's modulation ability. Since CAP is part of the vagus nerve and is directly innervated by the Medullary Visceral Zone (MVZ), we can reasonably concluded that pathological changes induced by MVZ's neuroinflammation should be responsible for CAP's dysfunction in modulating systemic inflammation in early sepsis. Methods: We conducted two independent septic experiments, the sepsis model rats were prepared by cecum ligation and puncture (CLP) method. In the first experiment, A total of 64 adult male Sprague-Dawley rats were included. Under the condition of sepsis and CAP's pharmacological activation or blockade, we investigated the MVZ's pathological changes, the functional state of key neurons including catecholaminergic and cholinergic neurons, key genes' expression such as Oligodendrocyte Transcription Factor 2 (Olig-2) mRNA, glial fibrillary acidic protein (GFAP) mRNA, and matrix metalloprotein (MMP) -9 mRNA, and CAP's activities reflected by HRV. The second experiment involved in 56 rats, through central anti-inflammation by feeding with 10 mg/ml minocycline sucrose solution as the only water source, or right vagus transection excepting for central anti-inflammation as a mean of the CAP's functional cancel, we confirmed that the neuroinflammation in MVZ affected systemic inflammation through CAP in sepsis. Results: In the first experiment, cholinergic and catecholaminergic neurons showed significant apoptosis with reduced expressions of TH, but the expression of CHAT remained relatively unaffected in MVZ in sepsis. HRV parameters representing the tone of the vagus nerve, such as SDNN, RMSSD, HF, SD1, and SD2, did not show significant differences among the three Septic Groups, although they all decreased significantly compared to the Control Group. The expressions of GFAP mRNA and MMP-9 mRNA were up-regulated, while the expression of Olig-2 mRNA was down-regulated in the Septic Groups. Intervention of CAP had a significant effect on cholinergic and catecholaminergic neurons' apoptosis, as well as the expressions of TH/CHAT and these key genes, but had little effect on HRV in sepsis. In the second experiment, the levels of TNF-α, IL-6, in serum and MVZ were significantly increased in sepsis. Central anti-inflammatory treatment reversed these changes. However, right vagotomy abolished the central anti-inflammatory effect. Conclusions: Our study uncovered that MVZ's neuroinflammation may play a crucial role in the uncontrolled systemic inflammation through inflammatory demyelination in MVZ, which disrupts CAP's modulation on the systemic inflammation in early sepsis.

EFFECTS ON DISTRIBUTION OF NADPH-d POSITIVE NEURONS AND Fos EXPRESSION IN THE MEDULLARY VISCERAL ZONE OF THE RAT FALLOWING CEREBRAL ISCHEMIA REPERFUSSION INJURY / 解剖学报

Objective To investigate the changes of distribution and expression of NADPH\|d positive neurons and Fos like immunoreactive(Fos\|LI) neurons in the rats medullary visceral zone(MVZ) following different time course after global cerebral ischemia reprefussion. Method Fifty adult SD rats were divided into three groups,normal control group( n =2),sham operation group( n =12) and ischemia group( n =36).The 4 brain vessel of ischemia group rats were occluded for 30?min,then reperfused for 2?h,4?h,8?h,12?h,24?h and 48?h respectively.The sections of medulla oblongata were divided into three series.The first and second series were reacted with NADPH\|d histochemical and anti\|Fos immunohistochemical methods respectively,and the third series were double stained with the other two methods mentioned above. Results NADPH\|d neurons and Fos\|LI neurons were distributed in bilateral MVZ(in NTS and VLM predominately).The number of NADPH\|d neurons and Fos\|LI neurons of ischemia group rats were significantly increased than that of control group at 2\|12?h.Approximately 15\|20 percent NADPH\|d/Fos\|LI double labeled neurons were found in MVZ.Conclusion\ The results indicated that NADPH\|d neurons and non\|NADPH\|d neurons could sensitively respond to brain ischemia reperfsion and might associate with regulation of stress and defence responese.

Fos expression in rat lumbarsacral spinal cord and medulla oblongata induced by chronic colonic inflammation / 中华消化杂志

Objective The aim of this study was to investigate Fos expression in rat lumbarsacral spinal cord and medulla oblongata induced by chronic colonic inflammation. Methods Twenty six male Sprague Dawley rats were divided into three groups: group 1, colonic inflammation was induced in sixteen rats by intraluminal instillation of trinitrobenzenesulfonic acid (TNBS); group 2, saline was instilled intraluminally in eight rats; group 3, no stimulation was given in 2 rats. After 3, 7, 14 and 28 days of instillation, lumbarsacral spinal cord and medulla oblongata were removed and processed for Fos immunohistochemical staining. Results Fos neurons induced by TNBS instillation were mainly distributed in deep laminae (laminae Ⅲ Ⅳ,Ⅴ Ⅵ) in spinal dorsal horn and in medullary visceral zone in medulla oblongata. The number of Fos cells in the spinal cord and medullary visceral zone was significantly higher in rats after 7 and 14 day of TNBS instillation compared with that of controls. After 28 days of TNBS instillation, the number of Fos neurons in the medullary visceral zone decreased and became comparable to that of control group. However, the number of Fos cells (54.1?16.3) in the spinal cord in some rats was still significantly higher than that of controls (12.2 ?2.6, P