Study on preparation and properties of silver alloy for Mongolian medicine acupuncture.

The Mongolian medical silver needles often encounter issues of bending, fracturing, and blunting in clinical applications. Similarly, Mongolian warm needles can cause burns on patients due to inaccurate temperature control. In this study, we developed an Ag85Cu15 alloy specifically for acupuncture needles based on material preparation. By incorporating appropriate amounts of Mn and Ti elements, we were able to enhance the mechanical properties and biocompatibility of the acupuncture needles. Compared to commercially available silver needles, this alloy exhibited a significant increase in microhardness up to 210.2 Hv0.2 and an improved tensile strength of 880.2 MPa. Furthermore, we designed a thermoelectric effect-based temperature measurement model for precise control of the warm needle's temperature, enhancing the therapeutic effectiveness of the treatment.

A computational framework of routine test data for the cost-effective chronic disease prediction.

Chronic diseases, because of insidious onset and long latent period, have become the major global disease burden. However, the current chronic disease diagnosis methods based on genetic markers or imaging analysis are challenging to promote completely due to high costs and cannot reach universality and popularization. This study analyzed massive data from routine blood and biochemical test of 32 448 patients and developed a novel framework for cost-effective chronic disease prediction with high accuracy (AUC 87.32%). Based on the best-performing XGBoost algorithm, 20 classification models were further constructed for 17 types of chronic diseases, including 9 types of cancers, 5 types of cardiovascular diseases and 3 types of mental illness. The highest accuracy of the model was 90.13% for cardia cancer, and the lowest was 76.38% for rectal cancer. The model interpretation with the SHAP algorithm showed that CREA, R-CV, GLU and NEUT% might be important indices to identify the most chronic diseases. PDW and R-CV are also discovered to be crucial indices in classifying the three types of chronic diseases (cardiovascular disease, cancer and mental illness). In addition, R-CV has a higher specificity for cancer, ALP for cardiovascular disease and GLU for mental illness. The association between chronic diseases was further revealed. At last, we build a user-friendly explainable machine-learning-based clinical decision support system (DisPioneer: http://bioinfor.imu.edu.cn/dispioneer) to assist in predicting, classifying and treating chronic diseases. This cost-effective work with simple blood tests will benefit more people and motivate clinical implementation and further investigation of chronic diseases prevention and surveillance program.

Analysis and identification of key anti-inflammatory molecules in Eerdun Wurile and exploration of their mechanism of action in microglia.

Ethnomedicine Eerdun Wurile (EW) can significantly promote poststroke neuro-recovery through modulation of microglia polarization. Fraction 4-6 (F4-6) isolated from EW via serial fractionation inhibits the expression of pro-inflammatory genes in LPS stimulated microglia. However, the key active molecules of F4-6 have not been identified. Herein, we identified alantolactone (Ala) and dehydrodiisoeugenol (Deh) as the active anti-inflammatory components of F4-6 by UPLC-qTof MS analysis. We confirmed that, F4-6, Ala, Deh and mixture of Ala and Deh (Mix) downregulate the expression of several pro-inflammatory genes including Ccl2, Cox2 and Il6 in LPS-treated microglia in a similar pattern. At the same time upregulate the expression of anti-inflammatory genes including Hmox1, Tgfß, Igf1 and Creb1. Moreover, the conditioned culture media obtained from F4-6 treated microglia significantly enhanced proliferation of N2a cells, and promoted neurite outgrowth possibly through upregulation of Nefh and Dlg4. Mechanistically, F4-6 strongly downregulated the expression of NF-κB p65, while also inhibiting the nuclear translocation of p65, leading to the suppression of transcription of pro-inflammatory genes initiated by NF-κB. Collectively, our data identified and quantified the key chemicals of EW and provide insights into the optimization of the herbal composition for neuroprotection.

Oral Delivery of SARS-CoV-2 DNA Vaccines Using Attenuated Salmonella typhimurium as a Carrier in Rat.

The 2019 novel coronavirus disease (COVID-19) is the disease that has been identified as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), but the prophylactic treatment of SARS-CoV-2 is still under investigation. The effective delivery of eukaryotic expression plasmids to the immune system's inductive cells constitutes an essential requirement for generating effective DNA vaccines. Here, we have explored the use of Salmonella typhimurium as vehicles to deliver expression plasmids orally. The attenuated Salmonella phoP was constructed by the one-step gene inactivation method, and plasmid-encoded the spike protein of SARS-CoV-2 was transform into the Salmonella phoP by electroporation. Western blot experiment was used for the detection of SARS-CoV-2 expression on 293T cells. Wistar rats were immunized orally with Salmonella that carried a eukaryotic expression plasmid once a week for three consecutive weeks. The ELISA was performed to measure the SARS-CoV-2 specific IgG at rat's serum samples. pSARS-CoV-2 can be successfully expression on 293T cells, and all immunized animals generated immunity against the SARS-CoV-2 spike protein, indicating that a Salmonella-based vaccine carrying the Spike gene can elicit SARS-CoV-2-specific secondary immune responses in rats. Oral delivery of SARS-CoV-2 DNA vaccines using attenuated Salmonella typhimurium may help develop a protective vaccine against SARS-CoV-2 infection.

Synergistic Regulation of Microglia Gene Expression by Natural Molecules in Herbal Medicine.

The activated microglia contribute to stroke-induced neuroinflammation by upregulating the expression of a pleura of genes that are characterized as either proinflammatory or anti-inflammatory. The natural products alantolactone (Ala) and dehydrodiisoeugenol (Deh) found in Inula helenium L. and Myristica fragrans Houtt., respectively, are regularly used in traditional herb medicine, which play anti-inflammatory and antioxidant roles via regulation of canonical pathways such as nuclear factor kappa B (NF-κB) in microglia and microphages. To illustrate the full spectra of gene expression alteration in microglia treated with Ala, Deh, and the mixture of Ala and Deh (denoted as Mix), we performed RNA-seq analysis of total RNA extracted from lipopolysaccharide- (LPS-) treated microglia subsequently exposed to Ala, Deh, and Mix. While both chemicals regulated the gene expression that facilitates an anti-inflammatory polarization, the mixture exerted some distinctive synergic regulatory effect, which differed from either of the chemicals alone. Our data provide important evidence for further research on the therapeutic mechanism of traditional medicine including Eerdun Wurile (EW).

Oral delivery of SARS-CoV-2 DNA vaccines using attenuated Salmonella typhimurium as a carrier in rat

The 2019 novel coronavirus disease (COVID-19) is the disease that has been identified as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), but the prophylactic treatment of SARS-CoV-2 is still under investigation. The effective delivery of eukaryotic expression plasmids to the immune systems inductive cells constitutes an essential requirement for the generation of effective DNA vaccines. Here, we have explored the use of Salmonella typhimurium as vehicles to deliver expression plasmids orally. Attenuated Salmonella phoP harboring eukaryotic expression plasmids that encoded spike protein of SARS-CoV-2 was administered orally to Wistar rats. Rats were immunized orally with Salmonella that carried a eukaryotic expression plasmid once a week for three consecutive weeks. The efficiency of the vaccination procedure was due to the transfer of the expression plasmid from the bacterial carrier to the mammalian host. Evidence for such an event could be obtained in vivo and in vitro. Our results showed that all immunized animals generated humoral immunity against the SARS-CoV-2 spike protein, indicating that a Salmonella-based vaccine carrying the Spike gene can elicit SARS-CoV-2-specific humoral immune responses in rats, and may be useful for the development of a protective vaccine against SARS-CoV-2 infection.

Bioactive components of ethnomedicine Eerdun Wurile regulate the transcription of pro-inflammatory cytokines in microglia.

ETHNOPHARMACOLOGICAL RELEVANCE: The traditional Mongolian medicine Eerdun Wurile (EW) has remarkable neural recovery effect, and has been playing a key role in the clinical treatment of neurological disorders including ischemic stroke in Inner Mongolia Autonomous Region of China. The preliminary pharmacological studies in animal suggested that EW regulates the expression of trophic factors in brain lesion and may also balance the polarization of activated microglia (Gaowa et al., 2018). AIM OF THE STUDY: The pool of leading bioactive chemicals underlying the therapeutic effects of EW has not been identified. Therefore, the mechanism of action of EW is poorly understood. This study was aimed to identify the major group of compounds that contribute to the inhibition of neuroinflammation during stroke recovery through regulation of microglia polarization. MATERIALS AND METHODS: The extracts of EW in different solvents were evaluated for their inhibitory ability of cytokine (IP-10) expression in LPS stimulated BV2 cells. The most effective extract (of petroleum ether extract) was further separated to 18 fractionations on a semi-preparative HPLC column, which were assess for the IP-10 down-regulation efficiency by RT-qPCR. The potent isolate was further fractionated in 12 fractions, which showed fewer peaks. The fraction 6 from this isolates, which remarkably down-regulates cytokines expression including IP-10, TNFα and IL-1ß, was analyzed on UPLC-qTOF MS. The key chemicals were measured for their cytokine inhibition in BV2 cells and mouse primary microglia. RESULTS: After two consecutive fractionating by preparative HPLC, petroleum ether extraction of EW gave 12 fractions with relatively distinctive chromatograms. A particular fraction (fraction 6) preserved the inhibitory effects on expression of pro-inflammatory cytokines including IP-10, TNFα, IL-1ß and iNOS. The result of UPLC-qTOF MS analysis showed that the fraction contains 21 chemicals including costunolide, alantolactone, myristicin and linolenic acid, which significantly down-regulate the expression of key pro-inflammatory cytokines in LPS stimulated BV2 cells as well as mouse primary microglia. CONCLUSION: Collectively our data suggest that the bioactive chemical pool which is responsible for the therapeutic effects of EW can be extracted in petroleum ether, and fractionated to a relatively small multiple components. Such components include known anti-inflammatory chemicals, which may contribute to the possible microglia polarization in brain lesion during the recovery of ischemic stroke.

Traditional Mongolian medicine Eerdun Wurile improves stroke recovery through regulation of gene expression in rat brain.

ETHNOPHARMACOLOGICAL RELEVANCE: Eerdun Wurile (EW) is one of the key Mongolian medicines for treatment of neurological and cardiological disorders. EW is ranked most regularly used Mongolian medicine in clinic. Components of EW which mainly originate from natural products are well defined and are unique to Mongolian medicine. AIM OF THE STUDY: Although the recipe of EW contains known neuroactive chemicals originated from plants, its mechanism of action has never been elucidated at molecular level. The objective of the present study is to explore the mechanism of neuroregenerative activity of EW by focusing on the regulation of gene expression in the brain of rat model of stroke. MATERIALS AND METHODS: Rat middle cerebral artery occlusion (MCAO) models were treated with EW for 15 days. Then, total RNAs from the cerebral cortex of rat MCAO models treated with either EW or control (saline) were extracted and analyzed by transcriptome sequencing. Differentially expressed genes were analyzed for their functions during the recovery of ischemic stroke. The expression level of significantly differentially expressed genes such as growth factors, microglia markers and secretive enzymes in the lesion was further validated by RT-qPCR and immunohistochemistry. RESULTS: Previously identified neuroactive compounds, such as geniposide (Yu et al., 2009), myristicin (Shin et al., 1988), costunolide (Okugawa et al., 1996), toosendanin (Shi and Chen, 1999) were detected in EW formulation. Bederson scale indicated that the treatment of rat MCAO models with EW showed significantly lowered neurological deficits (p < 0.01). The regional cerebral blood circulation was also remarkably higher in rat MCAO models treated with EW compared to the control group. A total of 186 genes were upregulated in the lesion of rat MCAO models treated with EW compared to control group. Among them, growth factors such as Igf1 (p < 0.05), Igf2 (p < 0.01), Grn (p < 0.01) were significantly upregulated in brain after treatment of rat MCAO models with EW. Meanwhile, greatly enhanced expression of microglia markers, as well as complementary components and secretive proteases were also detected. CONCLUSION: Our data collectively indicated that EW enhances expression of growth factors including Igf1 and Igf2 in neurons and microglia, and may stimulate microglia polarization in the brain. The consequences of such activity include stimulation of neuron growth, hydrolysis and clearance of cell debris at the lesion, as well as the angiogenesis.