Investigation of preclinical pharmacokinetics of N-demethylsinomenine, a potential novel analgesic candidate, using an UPLC-MS/MS quantification method.

N- Demethylsinomenine (NDSM), the in vivo demethylated metabolite of sinomenine, has exhibited antinociceptive efficacy against various pain models and may become a novel drug candidate for pain management. However, no reported analytical method for quantification of N- Demethylsinomenine in a biological matrix is currently available, and the pharmacokinetic properties of N- Demethylsinomenine are unknown. In the present study, an ultra-high performance liquid chromatography with tandem mass spectrometry (UPLC-MS/MS) method for quantification of N- Demethylsinomenine in rat plasma was developed and utilized to examine the preclinical pharmacokinetic profiles of N- Demethylsinomenine. The liquid-liquid extraction using ethyl acetate as the extractant was selected to treat rat plasma samples. The mixture of 25% aqueous phase (0.35% acetic acid-10 mM ammonium acetate buffer) and 75% organic phase (acetonitrile) was chosen as the mobile phases flowing on a ZORBAX C18 column to perform the chromatographic separation. After a 6-min rapid elution, NDSM and its internal standard (IS), metronidazole, were separated successfully. The ion pairs of 316/239 and 172/128 were captured for detecting N- Demethylsinomenine and IS, respectively, using multiple reaction monitoring (MRM) under a positive electrospray ionization (ESI) mode in this mass spectrometry analysis. The standard curve met linear requirements within the concentration range from 3 to 1000 ng/mL, and the lower limit of quantification (LLOQ) was 3 ng/mL. The method was evaluated regarding precision, accuracy, recovery, matrix effect, and stability, and all the results met the criteria presented in the guidelines for validation of biological analysis method. Then the pharmacokinetic profiles of N- Demethylsinomenine in rat plasma were characterized using this validated UPLC-MS/MS method. N- Demethylsinomenine exhibited the feature of linear pharmacokinetics after intravenous (i.v.) or intragastric (i.g.) administration in rats. After i. v. bolus at three dosage levels (0.5, 1, and 2 mg/kg), N- Demethylsinomenine showed the profiles of rapid elimination with mean half-life (T1/2Z) of 1.55-1.73 h, and extensive tissue distribution with volume of distribution (VZ) of 5.62-8.07 L/kg. After i. g. administration at three dosage levels (10, 20, and 40 mg/kg), N- Demethylsinomenine showed the consistent peak time (Tmax) of 3 h and the mean absolute bioavailability of N- Demethylsinomenine was 30.46%. These pharmacokinetics findings will aid in future drug development decisions of N- Demethylsinomenine as a potential candidate for pain analgesia.

Preclinical Pharmacokinetics and Bioavailability of Oxypeucedanin in Rats after Single Intravenous and Oral Administration.

Oxypeucedanin, a furanocoumarin extracted from many traditional Chinese herbal medicines, has a variety of pharmacological effects. However, the independent pharmacokinetic characteristics and bioavailability of this compound remains elusive. In this study, a rapid, sensitive, and selective method using ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC/MS/MS) was developed for evaluating the intravenous and oral pharmacokinetics of oxypeucedanin. After intravenous administration of oxypeucedanin (2.5, 5, and 10 mg/kg), and intragastric administration of oxypeucedanin (20 mg/kg), blood samples were collected periodically from the tail vein. The plasma concentration-time curves were plotted, and the pharmacokinetic parameters were calculated using a non-compartmental model analysis. After intravenous administration of oxypeucedanin (single dosing at 2.5, 5, and 10 mg/kg) to rats, the pharmacokinetics fit the linear kinetics characteristics, which showed that some parameters including average elimination half-life (T1/2Z of 0.61~0.66 h), mean residence time (MRT of 0.62~0.80 h), apparent volume of distribution (VZ of 4.98~7.50 L/kg), and systemic clearance (CLZ of 5.64~8.55 L/kg/h) are dose-independent and the area under concentration-time curve (AUC) increased in a dose-proportional manner. Single oral administration of oxypeucedanin (20 mg/kg) showed poor and slow absorption with the mean time to reach the peak concentration (Tmax) of 3.38 h, MRT of 5.86 h, T1/2Z of 2.94 h, and a mean absolute bioavailability of 10.26% in rats. These results provide critical information for a better understanding of the pharmacological effect of oxypeucedanin, which will facilitate its research and development.

N-Demethylsinomenine, an active metabolite of sinomenine, attenuates chronic neuropathic and inflammatory pain in mice.

Chronic pain is a significant public health problem that afflicts nearly 30% of the global population, but current pharmacotherapies are insufficient. Previous report indicated that N-demethylsinomenine, an active metabolite of sinomenine, is efficacious against postoperative pain. The present study investigated whether N-demethylsinomenine is effective for chronic painful conditions or whether repeated treatment alters its effect. Both chronic constriction injury (CCI) surgery and complete Freund's adjuvant (CFA) intraplantar injection induced significant and reliable mechanical allodynia at least for 7 days. Acute treatment with N-demethylsinomenine (10-40 mg/kg, i.p.) dose-dependently attenuated the mechanical allodynia both in CCI-induced neuropathic pain and CFA-induced inflammatory pain in mice. The potency of N-demethylsinomenine for reducing CFA-induced mechanical allodynia was slightly higher than sinomenine. During the period of repeated treatment, N-demethylsinomenine maintained its anti-allodynic effect against both neuropathic and inflammatory pain without producing carry-over effect. Pretreatment with bicuculline, a selective γ-aminobutyric acid type A (GABAA) receptor antagonist, almost completely blocked the anti-allodynia of N-demethylsinomenine (40 mg/kg) both in CCI and CFA-treated mice. Our findings indicated that N-demethylsinomenine exhibits GABAA receptor-mediated anti-allodynic effects in mouse models of neuropathic and inflammatory pain, suggesting it may be a useful novel pharmacotherapy for the control of chronic pain.

Lockdown Contained the Spread of 2019 Novel Coronavirus Disease in Huangshi City, China: Early Epidemiological Findings.

BACKGROUND: To control the spread of 2019 novel coronavirus disease (COVID-19), China sealed Wuhan on 23 January 2020 and soon expanded lockdown to 12 other cities in Hubei province. We aimed to describe the epidemiological characteristics in one of the cities and highlight the effect of current implemented lockdown and nonpharmaceutical interventions. METHODS: We retrieved data of reported cases in Huangshi and Wuhan from publicly available disease databases. Local epidemiological data on suspected or confirmed cases in Huangshi were collected through field investigation. Epidemic curves were constructed with data on reported and observed cases. RESULTS: The accumulated confirmed COVID-19 cases and fatality in Huangshi were reported to be 1015 and 3.74%, respectively, compared with 50006 and 5.08% in Wuhan until 27 March 2020. Right after 24 January, the epidemic curve based on observed cases in Huangshi became flattened. And 1 February 2020 was identified as the "turning point" as the epidemic in Huangshi faded soon afterward. COVID-19 epidemic was characterized by mild cases in Huangshi, accounting for 82.66% of total cases. Moreover, 50 asymptomatic infections were identified in adults and children. In addition, we found confirmed cases in 19 familial clusters and 21 healthcare workers, supporting interhuman transmission. CONCLUSIONS: Our study reported the temporal dynamics and characteristics of the COVID-19 epidemic in Huangshi city, China, across the unprecedented intervention. Such new epidemiological inference might provide further guidance on current lockdown measures in high-risk cities and, subsequently, help improve public health intervention strategies against the pandemic on the country and global levels.

Ligustilide prevents cognitive impairment and attenuates neurotoxicity in D-galactose induced aging mice brain.

Ligustilide (LIG) is a principal active ingredient of traditional Chinese medicine, Radix Angelica sinensis, which has versatile pharmacological activities including neuroprotection. Previous studies have demonstrated that LIG has beneficial effects on cognition deficits associated with cerebral damage or neurodegenerative disorders. In present study, we investigated the neuroprotective effect of LIG on cognitive impairment and neurotoxicity in the brain of aging mouse induced by d-galactose (d-gal). The aging model mice were induced by subcutaneous (S.C.) injection of d-gal once daily for 8 weeks and LIG (80 mg/kg) was simultaneously administered orally. The Morris water maze (MWM) test was used to assess the spatial learning and memory abilities. The activity of Na(+)-K(+)-ATPase and the content of lipid peroxidation product malondialdehyde (MDA) in brain were examined. The levels of glial fibrillary acidic protein (GFAP), growth-associated protein GAP-43, and cleaved caspase-3 in brain were also determined by immunohistochemistry. The MWM test showed that LIG administration markedly improved behavioral performance of d-gal treated mice. This action could be partly explained by the results that LIG reduced the level of MDA as well as increased the activity of Na(+)-K(+)-ATPase in the brain of d-gal induced aging mice. Moreover, LIG significantly raised the expression of GAP-43 and reduced cleaved caspase-3 and GFAP levels in the brain of d-gal treated mice. These results demonstrated that LIG improves d-gal-induced cognitive dysfunction and brain toxicity, which suggests that LIG may be developed as a new medicine for the treatment of aged-related conditions.

Chinese medicine approach in clinical practice for breast cancer survivors.

A Chinese medicine (CM) approach, designed based on the clinical experiences of the West Los Angeles Center for Traditional Chinese Medicine, is a practical path for assessing and treating breast cancer survivors. The approach consists of balancing the body with deciphering the cause of the patient's chief complaints by assessing and recognizing the six physiological dysfunctions that include gastrointestinal problems, sleeps problems, emotional imbalance, low body energy, menstrual change and pain. Multifaceted interventions are used for eliminating various CM pathologies based on identifying the basic CM patterns (syndromes) differentiation. Watching to assess the above two situations dynamically is used for outcome evaluation and predicting prognosis. Therefore this approach is called BMW. It can serve as a reference for CM clinical practice and integrative clinical care. It also can be used to simplify the clinical interpretation of CM and provide an easier way for CM doctors to communicate with Western medical doctors and patients. Additionally, it can be used as a guide for patients to assess their own symptoms for self-monitoring and self-care.

Patient education integrated with acupuncture for relief of cancer-related fatigue randomized controlled feasibility study.

BACKGROUND: Cancer-related fatigue (CRF) is a prominent clinical problem. There are calls for multi-modal interventions. METHODS: We assessed the feasibility of delivering patient education integrated with acupuncture for relief of CRF in a pilot randomized controlled trial (RCT) with breast cancer survivors using usual care as control. Social cognitive and integrative medicine theories guided integration of patient education with acupuncture into a coherent treatment protocol. The intervention consisted of two parts. First, patients were taught to improve self-care by optimizing exercise routines, improving nutrition, implementing some additional evidence-based cognitive behavioral techniques such as stress management in four weekly 50-minute sessions. Second, patients received eight weekly 50-minute acupuncture sessions. The pre-specified primary outcome, CRF, was assessed with the Brief Fatigue Inventory (BFI). Secondary outcomes included three dimensions of cognitive impairment assessed with the FACT-COGv2. RESULTS: Due to difficulties in recruitment, we tried several methods that led to the development of a tailored recruitment strategy: we enlisted oncologists into the core research team and recruited patients completing treatment from oncology waiting rooms. Compared to usual care control, the intervention was associated with a 2.38-point decline in fatigue as measured by the BFI (90% Confidence Interval from 0.586 to 5.014; p <0.10). Outcomes associated with cognitive dysfunction were not statistically significant. CONCLUSIONS: Patient education integrated with acupuncture had a very promising effect that warrants conducting a larger RCT to confirm findings. An effective recruitment strategy will be essential for the successful execution of a larger-scale trial. TRIAL REGISTRATION: NCT00646633.

Neuroprotective effect of astaxanthin on H(2)O(2)-induced neurotoxicity in vitro and on focal cerebral ischemia in vivo.

Astaxanthin (AST) is a powerful antioxidant that occurs naturally in a wide variety of living organisms. Much experimental evidence has proved that AST has the function of eliminating oxygen free radicals and can protect organisms from oxidative damage. The present study was carried out to further investigate the neuroprotective effect of AST on oxidative stress induced toxicity in primary culture of cortical neurons and on focal cerebral ischemia-reperfusion induced brain damage in rats. AST, over a concentration range of 250-1000nM, attenuated 50µM H(2)O(2)-induced cell viability loss. 500nM AST pretreatment significantly inhibited H(2)O(2)-induced apoptosis measured by Hoechst 33342 staining and restored the mitochondrial membrane potential (MMP) measured by a fluorescent dye, Rhodamine 123. In vivo, AST prevented cerebral ischemic injury induced by 2h middle cerebral artery occlusion (MCAO) and 24h reperfusion in rats. Pretreatment of AST intragastrically twice at 5h and 1h prior to ischemia dramatically diminished infarct volume and improved neurological deficit in a dose-dependent manner. Nissl staining showed that the neuronal injury was significantly improved by pretreatment of AST at 80mg/kg. Taken together, these results suggest that pretreatment with AST exhibits noticeable neuroprotection against brain damage induced by ischemia-reperfusion and the antioxidant activity of AST maybe partly responsible for it.

SiRNA targeting ICP4 attenuates HSV-1 replication / 病毒学报

HSV-1, a neurotropic virus, always leads to severe nervous symptoms. It is hard to completely eradicate the latent viruses after conventional therapy so that recurrence is inevitable. ICP is a key regulator for HSV replication and transcription that determines the cytolytic infection or latent state. In search of new anti-virus strategy targeting HSV-1ICP4, two pairs of siRNA were designed, and a recombinant eukaryotic lentiviral expression plasmid pLKO-puro(r)-hU6-siRNA was constructed. Vero cells were transfected with the designed siRNAs by Lipofectamine 2000 and four stable monoclonal cell lines were established by puromycin screening method. The ICP4 expression at mRNA level was detected with real-time PCR, and the HSV-1 replication was measured with TCID50 assay. SiRNA was shown as an effective way to inhibit the expression of ICP4 in monoclonal cell lines. Meanwhile, HSV-1 replication was significantly inhibited when ICP4 was shut down by siRNA. We conclude that siRNA targeting ICP4 attenuates HSV-1 replication. Further more, multi-site siRNAs show stronger inhibitory effect on viral replication, which may be an effective and feasible approach for biological anti-viral drugs.

[The protective effects of ginkgolide B and hypoxic preconditioning against acute hypoxia injury in mice].

AIM: To investigate the protective effects of ginkgolide B and hypoxic preconditioning against acute hypoxia injury in mice. METHODS: Ordinary pressure acute hypoxia model in mice was adopted to observe the ethology, the duration of the death and the degree of brain edema. Meanwhile the expression of RTP801 mRNA and erythropoietin (EPO) were measured by RT-PCR and Western blot, respectively. RESULTS: Ginkgolide B and hypoxic preconditioning could both prolong the survival time of hypoxia under ordinary pressure,and significantly decreased the degree of brain edema. Besides ginkgolide B and hypoxic preconditioning could both up-regulate the expression of RTP801mRNA and EPO. CONCLUSION: Ginkgolide B has the similar effect to hypoxic preconditioning against acute hypoxia. Both of these protective effects may be associated with the up-regulation of the expression of RTP801 mRNA and EPO.

Erxian tang--introduction of a Chinese herbal formula, clinical practice, and experimental studies.

Erxian Tang is a Chinese herbal formula developed for the treatment of menopausal syndrome in women. In the past 50 years, EXT has shown positive efficacy in the treatment of many chronic diseases in TCM, involving syndrome types of Shen yin-yang deficiency, yin-deficiency caused yang-flourishing, and disharmony of Chong-Ren meridian. Experimental studies have revealed that EXT has multiple pharmacological actions on such multiple targets as hypothalamus-pituitary-target gland axis, immune function and free radical metabolism, etc.

Introducing integrative East-West medicine to medical students and residents.

Over the last several years, many medical schools and residencies have introduced complementary and alternative medicine (CAM) into their curricula, prompting a discussion as to how CAM should be taught. In this paper, we share our experiences teaching medical trainees integrative East-West medicine, an approach to health and disease that brings together modern Western and Chinese medicine. A 2-week clinical rotation that is intimately tied to our busy clinical program is described in detail as we explore some of the challenges and opportunities involved in teaching a CAM-related field to medical trainees. We also demonstrate how such a clinical experience offers an opportunity to impart on our students a broad view of medicine and to discuss novel approaches to clinical problem-solving.