Yindan Xinnaotong capsule has been used for treating cardio-cerebrovascular diseases for several decades in China. Exercise training can protect against the development of atherosclerosis. The aim of the present study is to evaluate the joint effect of YXC and exercise on atherosclerosis in rats.

C-type lectin receptors (CLRs) play critical roles as pattern-recognition receptors (PRRs) for sensing Candida albicans infection, which can be life-threatening for immunocompromised individuals. Here we have shown that Dectin-3 (also called CLECSF8, MCL, or Clec4d), a previously uncharacterized CLR, recognized α-mannans on the surfaces of C. albicans hyphae and induced NF-κB activation. Mice with either blockade or genetically deleted Dectin-3 were highly susceptible to C. albicans infection. Dectin-3 constantly formed heterodimers with Dectin-2, a well-characterized CLR, for recognizing C. albicans hyphae. Compared to their respective homodimers, Dectin-3 and Dectin-2 heterodimers bound α-mannans more effectively, leading to potent inflammatory responses against fungal infections. Together, our study demonstrates that Dectin-3 forms a heterodimeric PRR with Dectin-2 for sensing fungal infection and suggests that different CLRs may form different hetero- and homodimers, which provide different sensitivity and diversity for host cells to detect various microbial infections.

The genomes of more than 50 organisms have now been manipulated due to rapid advancement of gene editing technology. One way to perform gene editing in the mouse using the CRISPR/CAS system, guide RNA (gRNA) and CAS9 mRNA transcribed in vitro are microinjected into fertilized eggs that are then allowed to develop to term. As a rule, gRNAs are tested first in tissue culture cells and the one with the highest locus-specific cleavage activity is chosen for microinjection. For cell transfections, gRNAs are typically expressed using the human U6 promoter (hU6). However, gRNAs for microinjection into zygotes are obtained by in vitro transcription from a T7 bacteriophage promoter in a separate plasmid vector. Here, we describe the design and construction of a combined U6T7 hybrid promoter from which the same gRNA sequence can be expressed. An expression vector containing such a hybrid promoter can now be used to generate gRNA for testing in mammalian cells as well as for microinjection purposes. The gRNAs expressed and transcribed from this vector are found to be functional in cells as well as in mice.

The adaptor protein CARD9 links detection of fungi by surface receptors to the activation of the NF-κB pathway. Mice deficient in CARD9 exhibit dysbiosis and are more susceptible to colitis. Here we examined the impact of Card9 deficiency in the development of colitis-associated colon cancer (CAC). Treatment of Card9-/- mice with AOM-DSS resulted in increased tumor loads as compared to WT mice and in the accumulation of myeloid-derived suppressor cells (MDSCs) in tumor tissue. The impaired fungicidal functions of Card9-/- macrophages led to increased fungal loads and variation in the overall composition of the intestinal mycobiota, with a notable increase in C. tropicalis. Bone marrow cells incubated with C. tropicalis exhibited MDSC features and suppressive functions. Fluconazole treatment suppressed CAC in Card9-/- mice and was associated with decreased MDSC accumulation. The frequency of MDSCs in tumor tissues of colon cancer patients correlated positively with fungal burden, pointing to the relevance of this regulatory axis in human disease.

Xueshuantong capsule (XST) is a patented traditional Chinese medicine used for the prevention and treatment of thrombosis. The molecular mechanism of anti-thrombotic effect of XST was investigated through the cross-talk among the platelets/leukocytes, endothelial cells (ECs), and flow shear stress. The Bioflux 1000 system was used to generate two levels of shear stress conditions: 0.1 and 0.9 Pa. Bioflux Metamorph microscopic imaging system was used to analyze the adhesion cell numbers. Protein expressions were detected by western blotting and flow cytometry. The flow-cytometry results showed that under 0.1 Pa flow, XST decreased ADP induced platelets CD62p surface expression in a concentration-dependent manner. Under 0.9 Pa flow, XST at a concentration of 0.15 g⋅L-1 reduced the platelets activation by 29.5%, and aspirin (ASA) showed no inhibitory effects. XST showed similar efficiency on monocytes adhesion both under 0.1 and 0.9 Pa flow conditions, and the inhibition rate was 30.2 and 28.3%, respectively. Under 0.9 Pa flow, the anti-adhesive effects of XST might be associated with the suppression of VE-cadherin and Cx43 in HUVECs. Blood flow not only acts as a drug transporter, but also exerts its effects to influence the pharmacodynamics of XST. Effects of XST on inhibiting platelets activation and suppressing platelets/leukocytes adhesion to injured ECs are not only concentration-dependent, but also shear stress-dependent. The mechanic forces combined with traditional Chinese medicine may be used as a precise treatment for cardiovascular diseases.

There are over 17,000 patients in the US waiting to receive liver transplants, and these numbers are increasing dramatically. Significant effort is being made to obtain functional hepatocytes and liver tissue that can for therapeutic use in patients. Blastocyst complementation is a challenging, innovative technology that could fundamentally change the future of organ transplantation. It requires the knockout (KO) of genes essential for cell or organ development in early stage host embryos followed by injection of donor pluripotent stem cells (PSCs) into host blastocysts to generate chimeric offspring in which progeny of the donor cells populate the open niche to develop functional tissues and organs.

Interactions between commensal fungi and gut immune system are critical for establishing colonic homeostasis. Here we found that mice deficient in Dectin-3 (Clec4d-/-), a C-type lectin receptor that senses fungal infection, were more susceptible to dextran sodium sulfate (DSS)-induced colitis compared with wild-type mice. The specific fungal burden of Candida (C.) tropicalis was markedly increased in the gut after DSS treatment in Clec4d-/- mice, and supplementation with C. tropicalis aggravated colitis only in Clec4d-/- mice, but not in wild-type controls. Mechanistically, Dectin-3 deficiency impairs phagocytic and fungicidal abilities of macrophages, and C. tropicalis-induced NF-κB activation and cytokine production. The conditioned media derived from Dectin-3-deficient macrophages were defective in promoting tissue repairing in colonic epithelial cells. Finally, anti-fungal therapy was effective in treating colitis in Clec4d-/- mice. These studies identified the role of Dectin-3 and its functional interaction with commensal fungi in intestinal immune system and regulation of colonic homeostasis.

We propose a Leaky-Wave Antenna (LWA) based on one-way yttrium-iron-garnet (YIG)-air-metal waveguide. We first analyze the dispersion of the LWA, showing the one-way feature and the radiation loss. Owing to the unique one-way dispersive property, the beam radiated from the LWA can have very narrow beam width, at the same time having large scanning angle. The main beam angle obtained by full-wave simulation is consistent with our theoretical prediction with the aid of the dispersion. For a given frequency, we can realize continuous beam scanning by varying the magnetic field, where the 3 dB beam width is much narrower than previously demonstrated. Our results pave a new way to realize continuous angle scanning at a fix frequency for modern communications.

XueShuanTong (XST) comprising therapeutically active ginsenosides, a lyophilized extract of Panax notoginseng roots, is extensively used in traditional Chinese medicine to treat ischemic heart and cerebrovascular diseases. Our recent study shows that treatment with XST inhibits shear-induced thrombosis formation but the underlying mechanism remained unclear. This study aimed to investigate the hypothesis that XST inhibited shear-induced platelet aggregation via targeting the mechanosensitive Ca2+-permeable Piezo1 channel by performing platelet aggregation assay, Ca2+ imaging and Western blotting analysis. Exposure to shear at physiologically (1,000-2000 s-1) and pathologically related rates (4,000-6,000 s-1) induced platelet aggregation that was inhibited by treatment with GsMTx-4. Exposure to shear evoked robust Ca2+ responses in platelets that were inhibited by treatment with GsMTx-4 and conversely enhanced by treatment with Yoda1. Treatment with XST at a clinically relevant concentration (0.15 g L-1) potently inhibited shear-induced Ca2+ responses and platelet aggregation, without altering vWF-mediated platelet adhesion and rolling. Exposure to shear, while resulting in no effect on the calpain-2 expression in platelets, induced calpain-2-mediated cleavage of talin1 protein, which is known to be critical for platelet activation. Shear-induced activation of calpain-2 and cleavage of talin1 were attenuated by treatment with XST. Taken together, our results suggest that XST inhibits shear-induced platelet aggregation via targeting the Piezo1 channel to prevent Piezo1-mediated Ca2+ signaling and downstream calpain-2 and talin1 signal pathway, thus providing novel insights into the mechanism of the therapeutic action of XST on platelet aggregation and thrombosis formation.

Spinocerebellar ataxia type 1 (SCA1) is a fatal neurodegenerative disease caused by an expanded polyglutamine tract in the widely expressed ATXN1 protein. To elucidate anatomical regions and cell types that underlie mutant ATXN1-induced disease phenotypes, we developed a floxed conditional knockout mouse model ( f-ATXN1 146Q/2Q ) having mouse Atxn1 coding exons replaced by human exons encoding 146 glutamines. F- ATXN1 146Q/2Q mice manifest SCA1-like phenotypes including motor and cognitive deficits, wasting, and decreased survival. CNS contributions to disease were revealed using ATXN1 146Q/2Q ; Nestin- Cre mice, that showed improved rotarod, open field and Barnes maze performances. Striatal contributions to motor deficits were examined using f- ATXN1 146Q/2Q ; Rgs9-Cre mice. Mice lacking striatal ATXN1 146Q/2Q had improved rotarod performance late in disease. Muscle contributions to disease were revealed in f- ATXN1 146Q/2Q ; ACTA1-Cre mice which lacked muscle pathology and kyphosis seen in f- ATXN1 146Q/2Q mice. Kyphosis was not improved in f-ATXN1 146Q/2Q ;Nestin-Cre mice. Thus, optimal SCA1 therapeutics will require targeting mutant ATXN1 toxic actions in multiple brain regions and muscle.

This research is the first to produce induced pluripotent stem cell-derived inner ear sensory neurons in the Neurog1+/- heterozygote mouse using blastocyst complementation. Additionally, this approach corrected non-sensory deficits associated with Neurog1 heterozygosity, indicating that complementation is specific to endogenous Neurog1 function. This work validates the use of blastocyst complementation as a tool to create novel insight into the function of developmental genes and highlights blastocyst complementation as a potential platform for generating chimeric inner ear cell types that can be transplanted into damaged inner ears to improve hearing.

Increasing evidence supports the hypothesis that inflammatory reactions serves an important function in the formation, progression and plaque rupture of atherosclerosis. Interleukin (IL)-1 primarily induces inflammation and is closely associated with the inflammatory environment and the formation of atherosclerosis. The present study aimed to establish an in vitro model for the evaluation of drug efficacy in the intervention of atherosclerosis from the inflammatory perspective, and to observe the anti-inflammatory effects of tanshinone IIA and andrographolide on atherosclerosis. The IL-1β-induced inflammation-activated endothelial cell (EC)-smooth muscle cell (SMC)-mononuclear cell (MC) co-culture model was established, based on the changes in a series of atherosclerosis-associated inflammatory markers secreted by ECs and SMCs. The expression of connexin in ECs, adhesion of MCs and changes in inflammatory signalling molecules were selected as evaluation indices for the inflammatory microenvironment of atherosclerosis. The use of this model revealed that tanshinone IIA exhibited significant efficacy against atherosclerosis and its inflammatory reactions. Inflammatory reactions were regarded as the primary mechanism underlying atherosclerosis. The established model simulated a series of relevant changes in the arterial wall under the inflammatory cytokines with oxidized low-density lipoprotein during the atherosclerotic process. The present study presented a reliable method for the identification of drugs with potential anti-inflammatory activity in atherosclerosis, for investigating the mechanisms of action, considering the improvement of the inflammatory state and the increase in plaque stability observed.

The TRPC5 proteins assemble to create calcium-permeable, non-selective, cationic channels. We sought novel modulators of these channels through studies of natural products.

Endothelial cells are sensitive to changes in both blood components and mechanical stimuli. Endothelial cells may undergo phenotypic changes, such as changes in adhesion protein expression, under different shear stress conditions. Such changes may impact platelet and monocyte adhesion to endothelial cells. This phenomenon is linked to chronic vascular inflammation and the development of atherosclerosis. In the present study, we investigated the effects of ginkgolide B on platelet and monocyte adhesion to human umbilical vein endothelial cells (HUVECs) under different conditions of laminar shear stress.

Objective. To investigate the absorption property of the representative hydrolyzable tannin, namely corilagin, and its hydrolysates gallic acid (GA) and ellagic acid (EA) from the Fructus Phyllanthi tannin fraction (PTF) in vitro. Methods. Caco-2 cells monolayer model was established. Influences of PTF on Caco-2 cells viability were detected with MTT assay. The transport across monolayers was examined for different time points, concentrations, and secretory directions. The inhibitors of P-glycoprotein (P-gp), multidrug resistance proteins (MRPs), organic anion transporting polypeptide (OATP) and sodium/glucose cotransporter 1 (SGLT1), and tight junction modulators were used to study the transport mechanism. LC-MS method was employed to quantify the absorption concentration. Results. The apparent permeability coefficient (Papp) values of the three compounds were below 1.0 × 10-6 cm/s. The absorption of corilagin and GA were much lower than their efflux, and the uptake of both compounds was increased in the presence of inhibitors of P-gp and MRPs. The absorption of EA was decreased in the company of OATP and SGLT1 inhibitors. Moreover, the transport of corilagin, GA, and EA was enhanced by tight junction modulators. Conclusion. These observations indicated that the three compounds in PTF were transported via passive diffusion combined with protein mediated transport. P-gp and MRPs might get involved in the transport of corilagin and GA. The absorption of EA could be attributed to OATP and SGLT1 protein.

Nemaline myopathy (NM) is one of the most common forms of congenital myopathy, and affects either fast myofibers, slow myofibers, or both. However, an animal model for congenital myopathy with fast-myofiber-specific atrophy is not available. Furthermore, mutations in the leiomodin-3 (LMOD3) gene have recently been identified in a group of individuals with NM. However, it is not clear how loss of LMOD3 leads to NM. Here, we report a mouse mutant in which the piggyBac (PB) transposon is inserted into the Lmod3 gene and disrupts its expression. Lmod3(PB/PB) mice show severe muscle weakness and postnatal growth retardation. Electron microscopy and immunofluorescence studies of the mutant skeletal muscles revealed the presence of nemaline bodies, a hallmark of NM, and disorganized sarcomeric structures. Interestingly, Lmod3 deficiency caused muscle atrophy specific to the fast fibers. Together, our results show that Lmod3 is required in the fast fibers for sarcomere integrity, and this study offers the first NM mouse model with muscle atrophy that is specific to fast fibers. This model could be a valuable resource for interrogating myopathy pathogenesis and developing therapeutics for NM as well as other pathophysiological conditions with preferential atrophy of fast fibers, including cancer cachexia and sarcopenia.

T cell receptor (TCR) transgenic mice represent an invaluable tool to study antigen-specific immune responses. In the pre-existing models, a monoclonal TCR is driven by a non-physiologic promoter and randomly integrated into the genome. Here, we create a highly efficient methodology to develop T cell receptor exchange (TRex) mice, in which TCRs, specific to the self/tumor antigen mesothelin (Msln), are integrated into the Trac locus, with concomitant Msln disruption to circumvent T cell tolerance. We show that high affinity TRex thymocytes undergo all sequential stages of maturation, express the exogenous TCR at DN4, require MHC class I for positive selection and undergo negative selection only when both Msln alleles are present. By comparison of TCRs with the same specificity but varying affinity, we show that Trac targeting improves functional sensitivity of a lower affinity TCR and confers resistance to T cell functional loss. By generating P14 TRex mice with the same specificity as the widely used LCMV-P14 TCR transgenic mouse, we demonstrate increased avidity of Trac-targeted TCRs over transgenic TCRs, while preserving physiologic T cell development. Together, our results support that the TRex methodology is an advanced tool to study physiological antigen-specific T cell behavior.

Glioblastoma is considered the most fatal malignant brain tumor that starts from the central nervous system (CNS), where the blood-brain barrier (BBB) remains the biggest challenge for active targeting of drugs in malignant brain tumor. Thereby, we have designed a paclitaxel PTX@ANG/FA-NPs hybrid novel nanodrug delivery system that can overcome the clinical BBB. The structural and morphological characterization of PTX@ANG/FA-NPs confirmed successful synthesis of nanomicelles with the size range of about 160 to 170 nm. The overall repressive effect of PTX@ANG/FA-NPs on human glioblastoma U251 cells was 1.2-times that of PTX alone. In vitro cellular uptake assay also demonstrated that the dual-targeted nanoparticles (NPs) were more easily taken up by glioblastoma U251 cells. Although the antiglioblastoma activity was confirmed by cell migration assay, apoptosis assay, and cellular uptake assay, the absorption was studied by in vivo fluorescence imaging and brain distribution. The synthesized PTX@ANG/FA-NPs probe significantly inhibited the migration of U251 within the cells and promoted the apoptosis process. Moreover, the RhB@ANG/FA-NPs and PTX@ANG/FA-NPs showed higher accumulating potential at sites of tumor BBB disruption. The novel nanodrug delivery system mediated enhanced distribution of drugs at the targeted site for therapeutics efficacies against glioblastomas across the BBB.

Spinocerebellar ataxia type 1 (SCA1) is a dominant trinucleotide repeat neurodegenerative disease characterized by motor dysfunction, cognitive impairment, and premature death. Degeneration of cerebellar Purkinje cells is a frequent and prominent pathological feature of SCA1. We previously showed that transport of ATXN1 to Purkinje cell nuclei is required for pathology, where mutant ATXN1 alters transcription. To examine the role of ATXN1 nuclear localization broadly in SCA1-like disease pathogenesis, CRISPR-Cas9 was used to develop a mouse with an amino acid alteration (K772T) in the nuclear localization sequence of the expanded ATXN1 protein. Characterization of these mice indicates that proper nuclear localization of mutant ATXN1 contributes to many disease-like phenotypes including motor dysfunction, cognitive deficits, and premature lethality. RNA sequencing analysis of genes with expression corrected to WT levels in Atxn1175QK772T/2Q mice indicates that transcriptomic aspects of SCA1 pathogenesis differ between the cerebellum, brainstem, cerebral cortex, hippocampus, and striatum.

Water-soluble tomato concentrate (WSTC), extracted from mature tomatoes, is the first health product in Europe that has been approved "to help maintain normal platelet activity to maintain healthy blood flow." We hypothesized that WSTC might exert an influence on blood flow shear stress-induced platelet aggregation (SIPA) and in turn maintains healthy blood flow. We used a microfluidic system to measure the effects of WSTC on SIPA in vitro. We also used the strenuous exercise rat model and the κ-carrageenan-induced rat tail thrombosis model to demonstrate the effects of WSTC on blood flow. WSTC significantly inhibited platelet aggregation at pathological high shear rate of 4,000 s-1 and 8,000 s-1 in vitro (P < 0.05 or P < 0.01). WSTC reduced the platelet adhesion rate and increased the rolling speed of platelets by inhibiting binding to Von Willebrand Factor (vWF) (P < 0.05 or P < 0.01). The oral administration of WSTC for 4 weeks in strenuous exercise rats alleviated hyper-reactivity of the platelets and led to a significant reduction in the plasma levels of catecholamine and IL-6. WSTC treatment also led to a reduction in black tail length, reduced blood flow pulse index (PI) and vascular resistance index (RI), and ameliorated local microcirculation perfusion in a rat model of thrombosis. WSTC exerted obvious inhibitory effects on the platelet aggregation induced by shear flow and alleviated the blood flow and microcirculation abnormities induced by an inflammatory reaction.