Site-Specific Covalent Immobilization of SMA-Stabilized ACE2 for SARS-CoV-2 Recognition and Drug Screening.

Membrane protein (MP)-based biomaterials have a wide range of applications in drug screening, antigen detection, and ligand-receptor interaction analysis. Traditional MP immobilization methods have the disadvantage of disordered protein immobilization orientation, leading to the shielded binding domain and unreliable binding pattern. Herein, we describe a site-specific covalent immobilization of MPs, which utilizes the styrene maleic acid (SMA) detergent-free extraction method of MPs as well as the covalent reaction between His-tag and divinyl sulfone (DVS). As an example, we covalently immobilized angiotensin-converting enzyme 2 (ACE2) on a cell membrane chromatography system (ACE2-His-SMALPs/CMC) in a site-specific manner and verified the specificity and stability of this system. This technique significantly improves the service life compared to the physisorption CMC column. The improved protein immobilization strategies of the ACE2-His-SMALPs/CMC system enable it to effectively recognize SARS-CoV-2 pseudoviral particles as well as detect viral particles in ambient air once combined with an aerosol collector; as a powerful ligand biosensor, the ACE2-His-SMALPs/CMC system was used to screen for compounds with anti-SARS-CoV-2 pseudovirus activity. In conclusion, the optimized MP immobilization strategy has been successfully applied to CMC technology, showing enhanced stability and sensitivity, which can provide an efficient and convenient membrane protein immobilization method for biomaterials.

5-Hydroxymethylfurfural multimers induce pseudo-allergic reaction through FcεRI at high doses.

Under acidic and high temperature conditions, 5-hydroxymethylfurfural (5-HMF) converted from sugar further produces dimers (Compound II) and trimers (Compound III). The polymers were less reported, and sensitization effect of them was reported in this study. Compounds II and III induced the local and systemic anaphylaxis effect in passive cutaneous anaphylaxis mice model and activated RBL-2H3 cell inducing [Ca2+ ] mobilization, resulting in the release of ß-hexosaminidase and histamine in vitro. The gene knockdown assay figured out that Compounds II and III induced degranulation through FcεRI. Further, Compounds II and III had a certain affinity with FcεRI by cell membrane chromatography and may combine on the "proline sandwich" structure indicated by molecular docking. All above suggested Compounds II and III can induce pseudo-allergic reaction through FcεRI in vivo and in vitro. Our work provides basic research to prove that the newly discovered 5-HMF transformants, Compounds II and III, induce pseudo-allergic reaction in vitro and in vivo through FcεRI, which is different pathway from 5-HMF. In foods with high sugar content, the sensitization of Compounds II and III needs more attention. In high-sugar foods and medicines, especially traditional Chinese medicine injections, the content of transformants needs to be detected.

P2RX7 in Dopaminergic Neurons of Ventral Periaqueductal Gray Mediates HTWP Acupuncture-Induced Consciousness in Traumatic Brain Injury.

The induction of a coma by traumatic brain injury (TBI) is a crucial factor for poor clinical prognoses. We report that acupuncture at the hand 12 Jing-Well points (HTWP) improved consciousness and neurologic function in TBI rats. Gene chip analyses showed that HTWP acupuncture mostly activated genes modulating neuronal projections (P2rx7, P2rx3, Trpv1, Tacr1, and Cacna1d), protein secretion (Exoc1, Exoc3l1, Fgb, and Fgr), and dopamine (DA) receptor D3 (Drd3) in the ventral periaqueductal gray (vPAG), among which the expression rate of P2rx7 was the most obviously increased. Acupuncture also increased the expression and excitability of DA and P2RX7 neurons, and the DA neurons expressed P2RX7, P2RX3, and TRPV1 in the vPAG. Intracerebroventricular administration of P2RX7, P2RX3, or TRPV1 antagonists blocked acupuncture-induced consciousness, and the subsequent injection of a P2RX7 antagonist into the vPAG nucleus also inhibited this effect. Our findings provide evidence that acupuncture alleviates TBI-induced comas via DA neurons expressing P2RX7 in the vPAG, so as to reveal the cellular and molecular mechanisms of the improvement of TBI clinical outcomes by HTWP acupuncture.

[Research advances in the clinical effect of bloodletting puncture at well-points in treatment of acute central nervous injury].

Bloodletting puncture at twelve well-points is a characteristic emergency therapy in traditional Chinese medicine. This article reviewed the research advances in the clinical effect of this therapy in the treatment of acute central nervous injury and its mechanism of action over the past 30 years, and it is found that this therapy can effectively improve disturbance of consciousness, neurological defects, and cerebral edema caused by stroke, traumatic brain injury, and carbon monoxide poisoning. The mechanism involves the improvement of cerebral blood flow and tissue oxygen supply, repair of the blood-brain barrier, and regulation of local ion balance. Well-designed clinical trials and in-depth research on biological mechanisms should be performed in future to promote and guide its clinical application.

Neurophysiology and Treatment of Disorders of Consciousness Induced by Traumatic Brain Injury: Orexin Signaling as a Potential Therapeutic Target.

BACKGROUND: Traumatic brain injury (TBI) can cause disorders of consciousness (DOC) by impairing the neuronal circuits of the ascending reticular activating system (ARAS) structures, including the hypothalamus, which are responsible for the maintenance of the wakefulness and awareness. However, the effectiveness of drugs targeting ARAS activation is still inadequate, and novel therapeutic modalities are urgently needed. METHODS: The goal of this work is to describe the neural loops of wakefulness, and explain how these elements participate in DOC, with emphasis on the identification of potential new therapeutic options for DOC induced by TBI. RESULTS: Hypothalamus has been identified as a sleep/wake center, and its anterior and posterior regions have diverse roles in the regulation of the sleep/wake function. In particular, the posterior hypothalamus (PH) possesses several types of neurons, including the orexin neurons in the lateral hypothalamus (LH) with widespread projections to other wakefulness-related regions of the brain. Orexins have been known to affect feeding and appetite, and recently their profound effect on sleep disorders and DOC has been identified. Orexin antagonists are used for the treatment of insomnia, and orexin agonists can be used for narcolepsy. Additionally, several studies demonstrated that the agonists of orexin might be effective in the treatment of DOC, providing novel therapeutic opportunities in this field. CONCLUSION: The hypothalamic-centered orexin has been adopted as the point of entry into the system of consciousness control, and modulators of orexin signaling opened several therapeutic opportunities for the treatment of DOC.