Histological and Ultrastructural Changes in Lungs One Year Post Asymptomatic or Mild SARS-CoV-2 Infection: Evaluating the Potential for Active Virus Presence / Cambios Histológicos y Ultraestructurales en los Pulmones un Año Después de una Infección Asintomática o Leve por SARS-CoV-2: Evaluación del Potencial de Presencia Activa del Virus

SUMMARY: Prior research on post-COVID-19 or long COVID primarily focused on the presence of SARS-CoV-2 mostly in symptomatic patients. This study aimed to investigate the persistence of SARS-CoV-2 after 1 year of asymptomatic or mild COVID-19. SARS-CoV-2 infected and control K18-hACE2 transgenic mice (n=25) were studied. Moderate and severe symptomatic subjects were sacrificed after eight days, while mild or asymptomatic mice were kept in BSL-III for twelve months. Analyses included general condition, histochemistry, immunohistochemistry, transmission electron microscopy, and qRT-PCR. Lungs from the twelve-month group showed thickening of alveolar walls, with some lungs exhibiting the recruitment of inflammatory cells, the presence of SARS- CoV-2 mRNA, immunopositivity for the SARS-CoV-2 spike protein, and TEM showed viruses (60-125 nm) within vesicles, indicating continued replication. Certain lung samples showed persistent SARS-CoV-2 presence in Club cells, endothelial cells, and macrophages. The eight-day group exhibited viral interstitial pneumonitis, SARS-CoV-2 immunopositivity, and mRNA. The eight-day hearts displayed viral mRNA, while the twelve-month hearts tested negative. Some asymptomatic twelve-month subjects presented reduced surfactant, basal membrane thickening, fibrosis, and mild autonomic nerve degeneration. In this study conducted on mice, findings indicate the potential for chronic persistence of SARS-CoV-2 in the lungs one year post initial mild or asymptomatic infection, which could suggest the possibility of recurrent episodes in similar human conditions. The observed thickening of alveolar walls and potential fibrotic areas in these mice may imply an increased risk of post-COVID fibrosis in humans. Furthermore, the presence of SARS-CoV-2-positive inflammatory cells in some asymptomatic murine cases could herald a progression toward ongoing inflammation and chronic lung disease in humans. Therefore, the necessity for further studies in human subjects and vigilant monitoring of high-risk human populations is underscored.

Investigaciones anteriores sobre COVID-19 o COVID prolongado se centraron principalmente en la presencia de SARS-CoV-2 principalmente en pacientes sintomáticos. Este estudio tuvo como objetivo investigar la persistencia del SARS-CoV-2 después de 1 año de COVID-19 asintomático o leve. Se estudiaron ratones transgénicos K18-hACE2 infectados con SARS-CoV-2 y de control (n=25). Los animales con síntomas moderados y graves se sacrificaron después de ocho días, mientras que los ratones con síntomas leves o asintomáticos se mantuvieron en BSL-III durante doce meses. Los análisis incluyeron estado general, histoquímica, inmunohistoquímica, microscopía electrónica de transmisión y qRT- PCR. Los pulmones del grupo de doce meses mostraron engrosamiento de las paredes alveolares, y algunos pulmones exhibieron reclutamiento de células inflamatorias, presencia de ARNm del SARS-CoV-2, inmunopositividad para la proteína de la espícula del SARS-CoV-2 y TEM mostró virus (60 -125 nm) dentro de las vesículas, lo que indica una replicación continua. Ciertas muestras de pulmón mostraron una presencia persistente de SARS- CoV-2 en exocrinocitos bronquiolares, células endoteliales y macrófagos. El grupo de ocho días presentó neumonitis intersticial viral, inmunopositividad al SARS-CoV-2 y ARNm. Los corazones de ocho días mostraron ARNm viral, mientras que los corazones de doce meses dieron negativo. Algunos animales asintomáticos de doce meses presentaron disminución del surfactante, engrosamiento de la membrana basal, fibrosis y degeneración leve del nervio autónomo. En este estudio realizado en ratones, los hallazgos indican la posibilidad de persistencia crónica del SARS-CoV-2 en los pulmones un año después de la infección inicial leve o asintomática, lo que podría sugerir la posibilidad de episodios recurrentes en condiciones humanas similares. El engrosamiento observado de las paredes alveolares y las posibles áreas fibróticas en estos ratones puede implicar un mayor riesgo de fibrosis post-COVID en humanos. Además, la presencia de células inflamatorias positivas para SARS- CoV-2 en algunos casos murinos asintomáticos podría presagiar una progresión hacia una inflamación continua y una enfermedad pulmonar crónica en humanos. Por lo tanto, se subraya la necesidad de realizar más estudios en seres humanos y realizar un seguimiento atento de las poblaciones humanas de alto riesgo.

Quercetin Alleviates Lipopolysaccharide-Induced Cardiac Inflammation via Inhibiting Autophagy and Programmed Cell Death / 生物医学与环境科学（英文）

OBJECTIVE@#The aim of this study is to explore the potential modulatory role of quercetin against Endotoxin or lipopolysaccharide (LPS) induced septic cardiac dysfunction.@*METHODS@#Specific pathogen-free chicken embryos ( n = 120) were allocated untreated control, phosphate buffer solution (PBS) vehicle, PBS with ethanol vehicle, LPS (500 ng/egg), LPS with quercetin treatment (10, 20, or 40 nmol/egg, respectively), Quercetin groups (10, 20, or 40 nmol/egg). Fifteen-day-old embryonated eggs were inoculated with abovementioned solutions via the allantoic cavity. At embryonic day 19, the hearts of the embryos were collected for histopathological examination, RNA extraction, real-time polymerase chain reaction, immunohistochemical investigations, and Western blotting.@*RESULTS@#They demonstrated that the heart presented inflammatory responses after LPS induction. The LPS-induced higher mRNA expressions of inflammation-related factors (TLR4, TNFα, MYD88, NF-κB1, IFNγ, IL-1β, IL-8, IL-6, IL-10, p38, MMP3, and MMP9) were blocked by quercetin with three dosages. Quercetin significantly decreased immunopositivity to TLR4 and MMP9 in the treatment group when compared with the LPS group. Quercetin significantly decreased protein expressions of TLR4, IFNγ, MMP3, and MMP9 when compared with the LPS group. Quercetin treatment prevented LPS-induced increase in the mRNA expression of Claudin 1 and ZO-1, and significantly decreased protein expression of claudin 1 when compared with the LPS group. Quercetin significantly downregulated autophagy-related gene expressions (PPARα, SGLT1, APOA4, AMPKα1, AMPKα2, ATG5, ATG7, Beclin-1, and LC3B) and programmed cell death (Fas, Bcl-2, CASP1, CASP12, CASP3, and RIPK1) after LPS induction. Quercetin significantly decreased immunopositivity to APOA4, AMPKα2, and LC3-II/LC3-I in the treatment group when compared with the LPS group. Quercetin significantly decreased protein expressions of AMPKα1, LC3-I, and LC3-II. Quercetin significantly decreased the protein expression to CASP1 and CASP3 by immunohistochemical investigation or Western blotting in treatment group when compared with LPS group.@*CONCLUSION@#Quercetin alleviates cardiac inflammation induced by LPS through modulating autophagy, programmed cell death, and myocardiocytes permeability.

Polysaccharide of Alocasia cucullata Exerts Antitumor Effect by Regulating Bcl-2, Caspase-3 and ERK1/2 Expressions during Long-Time Administration / 中国结合医学杂志

OBJECTIVE@#To study the in vitro and in vivo antitumor effects of the polysaccharide of Alocasia cucullata (PAC) and the underlying mechanism.@*METHODS@#B16F10 and 4T1 cells were cultured with PAC of 40 µg/mL, and PAC was withdrawn after 40 days of administration. The cell viability was detected by cell counting kit-8. The expression of Bcl-2 and Caspase-3 proteins were detected by Western blot and the expressions of ERK1/2 mRNA were detected by quantitative real-time polymerase chain reaction (qRT-PCR). A mouse melanoma model was established to study the effect of PAC during long-time administration. Mice were divided into 3 treatment groups: control group treated with saline water, positive control group (LNT group) treated with lentinan at 100 mg/(kg·d), and PAC group treated with PAC at 120 mg/(kg·d). The pathological changes of tumor tissues were observed by hematoxylin-eosin staining. The apoptosis of tumor tissues was detected by TUNEL staining. Bcl-2 and Caspase-3 protein expressions were detected by immunohistochemistry, and the expressions of ERK1/2, JNK1 and p38 mRNA were detected by qRT-PCR.@*RESULTS@#In vitro, no strong inhibitory effects of PAC were found in various tumor cells after 48 or 72 h of administration. Interestingly however, after 40 days of cultivation under PAC, an inhibitory effect on B16F10 cells was found. Correspondingly, the long-time administration of PAC led to downregulation of Bcl-2 protein (P<0.05), up-regulation of Caspase-3 protein (P<0.05) and ERK1 mRNA (P<0.05) in B16F10 cells. The above results were verified by in vivo experiments. In addition, viability of B16F10 cells under long-time administration culture in vitro decreased after drug withdrawal, and similar results were also observed in 4T1 cells.@*CONCLUSIONS@#Long-time administration of PAC can significantly inhibit viability and promote apoptosis of tumor cells, and had obvious antitumor effect in tumor-bearing mice.

Hydroxysafflor Yellow A Promotes HaCaT Cell Proliferation and Migration by Regulating HBEGF/EGFR and PI3K/AKT Pathways and Circ_0084443 / 中国结合医学杂志

OBJECTIVE@#To investigate the effect and possible mechanism of hydroxysafflor yellow A (HSYA) on human immortalized keratinocyte cell proliferation and migration.@*METHODS@#HaCaT cells were treated with HSYA. Cell proliferation was detected by the cell counting kit-8 assay, and cell migration was measured using wound healing assay and Transwell migration assay. The mRNA and protein expression levels of heparin-binding epidermal growth factor (EGF)-like growth factor (HBEGF), EGF receptor (EGFR), phosphatidylinositol 3-kinase (PI3K), protein kinase B (AKT), mammalian target of rapamycin (mTOR), and hypoxia-inducible factor-1α (HIF-1α) were detected by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot, respectively. Circ_0084443-overexpressing HaCaT cells and empty plasmid HaCaT cells were constructed using the lentiviral stable transfection and treated with HSYA. The expression of circ_0084443 was detected by qRT-PCR.@*RESULTS@#HSYA (800 µmol/L) significantly promoted HaCaT cell proliferation and migration (P<0.05 or P<0.01). It also increased the mRNA and protein expression levels of HBEGF, EGFR, PI3K, AKT, mTOR and HIF-1α, and increased the phosphorylation levels of PI3K and AKT (P<0.05 or P<0.01). Furthermore, HSYA promoted HaCaT cell proliferation and migration via the HBEGF/EGFR and PI3K/AKT/mTOR signaling pathways (P<0.01). Circ_0084443 attenuated the mRNA expression levels of HBEGF, EGFR, PI3K, AKT, mTOR and HIF-1α (P<0.05). HSYA inhibited the circ_0084443 expression, further antagonized the inhibition of circ_0084443 on HBEGF, EGFR, PI3K, AKT, mTOR and HIF-1α, and promoted the proliferation of circ_0084443-overexpressing HaCaT cells (P<0.05 or P<0.01). However, HSYA could not influence the inhibitory effect of circ_0084443 on HaCaT cell migration (P>0.05).@*CONCLUSION@#HSYA played an accelerative role in HaCaT cell proliferation and migration, which may be attributable to activating HBEGF/EGFR and PI3K/AKT signaling pathways, and had a particular inhibitory effect on the keratinocyte negative regulator circ_0084443.

Effects of canagliflozin on amino acid metabolism in atherosclerotic mice / 中华心血管病杂志

Objective: To explore the possible anti-atherosclerotic mechanisms of glucose co-transporter-2 inhibitor canagliflozin. Methods: ApoE-/-mice fed on Western diet were randomly assigned into the model group (n=10) and the canagliflozin group (n=10). C57BL/6J mice fed on normal diet were chosen as the control group (n=10). Mice in the canagliflozin group were gavaged with canagliflozin for 14 weeks. The presence and severity of atherosclerosis were evaluated with HE and oil red O stainings in aortic root section slices. PCR assay was performed to determine the mRNA expression levels of nitric oxide synthase. Hepatic transcriptome analysis and hepatic amino acid detection were conducted using RNA-seq and targeted LC-MS, respectively. Results: HE staining and oil red O staining of the aortic root showed that AS models were successfully established in ApoE-/-mice fed on Western diet for 14 weeks. Canagliflozin alleviated the severity of atherosclerosis in pathology. Hepatic transcriptome analysis indicated that canagliflozin impacted on amino acid metabolism, especially arginine synthesis in ApoE-/-mice. Targeted metabolomics analysis of amino acids showed that canagliflozin reduced hepatic levels of L-serine, L-aspartic acid, tyrosine, L-hydroxyproline, and L-citrulline, but raised the hepatic level of L-arginine. Compared to the model group, the canagliflozin group exhibited higher serum arginine and nitric oxide levels as well as elevated nitric oxide mRNA expression in aortic tissues (P<0.05). Conclusion: Canagliflozin regulated the amino acid metabolism, reduced the levels of glucogenic amino acids,and promoted the synthesis of arginine in atherosclerotic mice.

Effects of canagliflozin on amino acid metabolism in atherosclerotic mice / 中华心血管病杂志

Objective: To explore the possible anti-atherosclerotic mechanisms of glucose co-transporter-2 inhibitor canagliflozin. Methods: ApoE-/-mice fed on Western diet were randomly assigned into the model group (n=10) and the canagliflozin group (n=10). C57BL/6J mice fed on normal diet were chosen as the control group (n=10). Mice in the canagliflozin group were gavaged with canagliflozin for 14 weeks. The presence and severity of atherosclerosis were evaluated with HE and oil red O stainings in aortic root section slices. PCR assay was performed to determine the mRNA expression levels of nitric oxide synthase. Hepatic transcriptome analysis and hepatic amino acid detection were conducted using RNA-seq and targeted LC-MS, respectively. Results: HE staining and oil red O staining of the aortic root showed that AS models were successfully established in ApoE-/-mice fed on Western diet for 14 weeks. Canagliflozin alleviated the severity of atherosclerosis in pathology. Hepatic transcriptome analysis indicated that canagliflozin impacted on amino acid metabolism, especially arginine synthesis in ApoE-/-mice. Targeted metabolomics analysis of amino acids showed that canagliflozin reduced hepatic levels of L-serine, L-aspartic acid, tyrosine, L-hydroxyproline, and L-citrulline, but raised the hepatic level of L-arginine. Compared to the model group, the canagliflozin group exhibited higher serum arginine and nitric oxide levels as well as elevated nitric oxide mRNA expression in aortic tissues (P<0.05). Conclusion: Canagliflozin regulated the amino acid metabolism, reduced the levels of glucogenic amino acids,and promoted the synthesis of arginine in atherosclerotic mice.

Effect of Survivin Down-Regulation by Egr1-Survivin shRNA Combined with Radiotherapy on Radiosensitivity of Esophageal Squamous Carcinoma / Efecto de la Regulación Negativa de Survivina por shRNA de Egr1-Survivina Combinado con Radioterapia sobre la Radiosensibilidad del Carcinoma Escamoso de Esófago

SUMMARY: This study is to investigate the effect of survivin down-regulation by Egr1-survivin shRNA combined with radiotherapy on the apoptosis and radiosensitivity of esophageal squamous cell carcinoma ECA109 and KYSE150 cells. ECA109 and KYSE150 cells were transfected with Egr1-survivin shRNA, and then treated with radiotherapy. After 24 h, the mRNA and protein levels of Egr1-survivin were detected by qPCR and Western-Blot. Cell cycle and apoptosis were detected by flow cytometry. Western blot also detected levels of cleavaged Caspase 3 and Caspase 9. YM155 was used as a positive control to inhibit survivin expression. The levels of survivin mRNA and protein in ECA109 and KYSE150 cells treated with Egr1-survivin shRNA combined with radiotherapy were significantly lower than those of the blank control group, the empty vector control group, and, the YM155 + radiotherapy group (P<0.05). Meanwhile, after survivin down-regulation, the ratio of G2 to S phase of ECA109 and KYSE150 cells increased significantly, leading to significant G2 and S phase arrest. Additionally, apoptosis of ECA109 and KYSE150 cells increased significantly (P <0.01). Further, protein levels of cleavaged Caspase 3 and Caspase 9 significantly increased in Egr1-survivin shRNA combined with radiotherapy group. Egr1-survivin shRNA combined with radiotherapy can down-regulate survivin expression, which further increases the apoptosis, and enhances the radiosensitivity of ECA109 and KYSE150 cells.

Este estudio tuvo como objetivo investigar el efecto de la regulación negativa de survivina por el shRNA de Egr1-survivina combinado con radioterapia sobre la apoptosis y la radiosensibilidad del carcinoma de células escamosas de esófago Células ECA109 y KYSE150. Las células ECA109 y KYSE150 se transfectaron con shRNA de survivina Egr1 y luego se trataron con radioterapia. Después de 24 h, los niveles de ARNm y proteína de Egr1-survivina se detectaron mediante qPCR y Western-Blot. El ciclo celular y la apoptosis se detectaron mediante citometría de flujo. La transferencia Western también detectó niveles de Caspasa 3 y Caspasa 9 escindidas. Se usó YM155 como control positivo para inhibir la expresión de survivina. Los niveles de ARNm y proteína de survivina en células ECA109 y KYSE150 tratadas con shRNA de survivina Egr1 combinado con radioterapia fueron significativamente más bajos que los del grupo control en blanco, el grupo control de vector vacío y el grupo de radioterapia YM155 + (P <0,05). Mientras tanto, después de la regulación negativa de survivina, la proporción entre las fases G2 y S de las células ECA109 y KYSE150 aumentó significativamente, lo que llevó a una detención significativa de las fases G2 y S. Además, la apoptosis de las células ECA109 y KYSE150 aumentó significativamente (P <0,01). Además, los niveles de proteína de Caspasa 3 y Caspasa 9 escindidas aumentaron significativamente en el shRNA de Egr1- survivina combinado con el grupo de radioterapia. El shRNA de survivina de Egr1 combinado con radioterapia puede regular negativamente la expresión de survivina, lo que aumenta aún más la apoptosis y mejora la radiosensibilidad de las células ECA109 y KYSE150.

TTYH3 is a prognostic biomarker and associates with immune cell infiltrations of lung adenocarcinoma / TTYH3 es un biomarcador pronóstico y se asocia con infiltraciones de células inmunitarias del adenocarcinoma de pulmón

SUMMARY: We investigated Tweety Family Member 3 (TTYH3) level in lung adenocarcinoma (LUAD) and its relationship with immune infiltration in tumors by bioinformatics. Differential expressions of TTYH3 in lung cancer were analyzed with Oncomine, TIMER, GEO, UALCAN and HPA. Relationship of TTYH3 mRNA/protein levels with clinical parameters was analyzed by UALCAN. Co-expressed genes of TTYH3 in LUAD were analyzed using Cbioportal. Its relationship with LUAD prognosis was analyzed by Kaplan-Meier plotter. GO and KEGG analysis were performed. Correlation between TTYH3 and tumor immune infiltration were tested by TIMER, TISIDB and GEPIA. We found that TTYH3 was significantly increased in LUAD tissues. TTYH3 high expression was closely related to poor overall survival, post progression survival and first progression in LUAD patients. TTYH3 mRNA/protein levels were significantly associated with multiple pathways. Specifically, TTYH3 up-regulation was mostly related to biological regulation, metabolic process, protein blinding, extracellular matrix organization and pathways in cancer. Moreover, TTYH3 was positively associated with immune cell infiltration in LUAD. Finally, TTYH3 was highly expressed in LUAD as revealed by meta-analysis. TTYH3 is closely related to the prognosis of LUAD and immune cell infiltration, and it can be used as a prognostic biomarker for LUAD and immune infiltration.

Investigamos por bioinformática el nivel de Tweety Family Member 3 (TTYH3) con adenocarcinoma de pulmón (LUAD) y su relación con la infiltración inmune en tumores. Las expresiones diferenciales de TTYH3 en cáncer de pulmón se analizaron con Oncomine, TIMER, GEO, UALCAN y HPA. Con UALCAN se analizó la relación de los niveles de ARNm/proteína de TTYH3 con los parámetros clínicos. Los genes coexpresados de TTYH3 en LUAD se analizaron utilizando Cbioportal. Su relación con el pronóstico LUAD se analizó mediante plotter de Kaplan- Meier. Se realizaron análisis GO y KEGG. TIMER, TISIDB y GEPIA probaron la correlación entre TTYH3 y la infiltración inmune tumoral. Encontramos que TTYH3 aumentó significativamente en los tejidos LUAD. La alta expresión de TTYH3 estuvo estrechamente relacionada con una supervivencia general deficiente, supervivencia posterior a la progresión y primera progresión en pacientes con LUAD. Los niveles de ARNm/ proteína de TTYH3 se asociaron significativamente con múltiples vías. Específicamente, la regulación positiva de TTYH3 se relacionó principalmente con la regulación biológica, el proceso metabólico, el cegamiento de proteínas, la organización de la matriz extracelular y las vías en el cáncer. Además, TTYH3 se asoció positivamente con la infiltración de células inmunitarias en LUAD. Finalmente, TTYH3 se expresó altamente en LUAD como lo reveló el metanálisis. TTYH3 está estrechamente relacionado con el pronóstico de LUAD y la infiltración de células inmunitarias, y se puede utilizar como biomarcador pronóstico para LUAD y la infiltración de células inmunitarias.

Effect of acupuncture at "Zhibian" (BL 54) through "Shuidao" (ST 28) on the expression of apoptosis-related factors in rats with premature ovarian insufficiency based on oxidative stress / 中国针灸

OBJECTIVE@#To explore the possible mechanism of acupuncture at "Zhibian" (BL 54) through "Shuidao" (ST 28) on premature ovarian insufficiency (POI) from the perspective of oxidative stress.@*METHODS@#Sixty female SD rats were randomly divided into a blank group, a model group, a sham acupuncture group, a medication group, and an acupuncture group, 12 rats in each group. Except the blank group, the rats in the remaining groups were intraperitoneally injected with cyclophosphamide to establish the POI model. After the model was successfully established, the rats in the acupuncture group were treated with acupuncture at "Zhibian" (BL 54) through "Shuidao" (ST 28), with a depth of about 12 mm, and the needle was retained for 30 min; the acupuncture was given once a day, for a total of 4 weeks. The rats in the sham acupuncture group were treated with blunt-head needle to tap the skin surface of "Zhibian" (BL 54), without penetrating the skin, once a day for 4 weeks. The rats in the medication group were treated with estradiol valerate by gastric gavage for 4 weeks. After the intervention, the level of reactive oxygen species (ROS) in the ovarian tissue was detected by fluorescence probe; the expression of c-Jun N-terminal kinase (JNK), forkhead box O1 (FoxO1), tumor suppressor gene protein 53 (p53) and p53 up-regulated modulator of apoptosis (Puma) mRNA and protein in ovarian tissue were detected by real-time fluorescence quantitative PCR and Western blot.@*RESULTS@#Compared with the blank group, the level of ROS and the expression of JNK mRNA, p-JNK protein, FoxO1, p53, Puma mRNA and protein in the ovarian tissue in the model group were increased (P<0.01). Compared with the model group, the level of ROS and the expression of p-JNK protein, FoxO1, p53, Puma mRNA and protein in the ovarian tissue in the sham acupuncture group were slightly reduced, but the difference was not statistically significant (P>0.05). The level of ROS and the expression of JNK mRNA, p-JNK protein, FoxO1, p53, Puma mRNA and protein in the ovarian tissue in the acupuncture group and the medication group were reduced (P<0.01).@*CONCLUSION@#Acupuncture at "Zhibian" (BL 54) through "Shuidao" (ST 28) could improve the level of oxidative stress, down-regulate the expression of apoptosis-related factors JNK, FoxO1, p53 and Puma induced by oxidative stress, and inhibit the premature failure of ovarian reserve function caused by apoptosis of ovarian granulosa cells in POI rats.

Effects of electroacupuncture on NLRP3/Caspase-1/GSDMD axis and neurological function in rats with cerebral ischemic reperfusion / 中国针灸

OBJECTIVE@#To investigate the neuroprotective effect of electroacupuncture (EA) at "Quchi" (LI 11) and "Zusanli" (ST 36) in the rats with cerebral ischemic reperfusion and the potential mechanism of microglia pyroptosis.@*METHODS@#Sixty SD rats were randomly divided into a sham-operation group, a model group and an EA group, with 20 rats in each group. The Zea Longa method was employed to establish the rat model of the middle cerebral artery occlusion and reperfusion (MACO/R) in the left brain. In the EA group, since the 2nd day of modeling, EA was given at "Quchi" (LI 11) and "Zusanli" (ST 36) of right side with disperse-dense wave, 4 Hz/20 Hz in frequency and 0.2 mA in current intensity, 30 min each time, once a day for lasting 7 consecutive days. The reduction rate of cerebral blood flow was measured with laser Doppler flowmetry during operation. The neurological function of rats was observed using Zea Longa neurobehavioral score. The cerebral infarction volume was detected by TTC staining method. The microglia positive expression in the ischemic side of the cortex was detected with the immunofluorescence method. Under transmission electron microscope, the ultrastructure of cell in the ischemic cortex was observed. The mRNA expression levels of nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3), apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC), cysteinyl aspartate specific proteinase-1 (Caspase-1) and gasdermin D (GSDMD) in the ischemic cortex were detected using real-time PCR.@*RESULTS@#Compared with the sham-operation group, in the model group, the reduction rate of cerebral blood flow was increased during operation (P<0.001); Zea Longa neurobehavional score and the percentage of cerebral infarction volume were increased (P<0.001), the numbers of M1-type microglia marked by CD68+ and M2-type microglia marked by TMEM119+ were elevated in the ischemic cortex (P<0.001), the mRNA expression of NLRP3, ASC, Caspase-1 and GSDMD was increased (P<0.001, P<0.01); the cytomembrane structure was destroyed, with more cell membrane pores formed in the ischemic cortex. Compared with the model group, after intervention, Zea Longa neurobehavioral score and the percentage of cerebral infarction volume were reduced (P<0.05), the number of M1-type microglia marked by CD68+ was reduced (P<0.05) and the number of M2-type microglia marked by TMEM119+ was increased (P<0.05); and the mRNA expression of NLRP3, ASC, Caspase-1 and GSDMD was decreased (P<0.01, P<0.05) in the EA group. Even though the cytomembrane structure was incomplete, there were less membrane pores presented in the ischemic cortex in the EA group after intervention.@*CONCLUSION@#The intervention with EA attenuates the neurological dysfunction and reduces the volume of cerebral infarction in the rats with cerebral ischemic reperfusion. The underlying mechanism is related to the inhibition of microglia pyroptosis through modulating NLRP3/Caspase-1/GSDMD axis.

Effects of electroacupuncture pretreatment on GABAA receptor of fastigial nucleus and sympathetic nerve activity in rats with myocardial ischemia reperfusion injury / 中国针灸

OBJECTIVE@#To observe the effects of electroacupuncture (EA) pretreatment on cardiac function, sympathetic nerve activity, indexes of myocardial injury and GABAA receptor in fastigial nucleus in rats with myocardial ischemia reperfusion injury (MIRI), and to explore the neuroregulatory mechanism of EA pretreatment in improving MIRI.@*METHODS@#A total of 60 male SD rats were randomly divided into a sham operation group, a model group, an EA group, an agonist group and an agonist+EA group, 12 rats in each group. The MIRI model was established by ligation of the left anterior descending coronary artery. EA was applied at bilateral "Shenmen" (HT 7) and "Tongli" (HT 5) in the EA group and the agonist+EA group, with continuous wave, in frequency of 2 Hz and intensity of 1 mA, 30 min each time, once a day for 7 consecutive days. After intervention, the MIRI model was established. In the agonist group, the muscone (agonist of GABAA receptor, 1 g/L) was injected in fastigial nucleus for 7 consecutive days before modeling, 150 μL each time, once a day. In the agonist+EA group, the muscone was injected in fastigial nucleus 30 min before EA intervention. The data of electrocardiogram was collected by PowerLab standard Ⅱ lead, and ST segment displacement and heart rate variability (HRV) were analyzed; the serum levels of norepinephrine (NE), creatine kinase isoenzyme MB (CK-MB) and cardiac troponin I (cTnI) were detected by ELISA; the myocardial infarction area was measured by TTC staining; the morphology of myocardial tissue was observed by HE staining; the positive expression and mRNA expression of GABAA receptor in fastigial nucleus were detected by immunohistochemistry and real-time PCR.@*RESULTS@#Compared with the sham operation group, in the model group, ST segment displacement and ratio of low frequency to high frequency (LF/HF) of HRV were increased (P<0.01), HRV frequency domain analysis showed enhanced sympathetic nerve excitability, the serum levels of NE, CK-MB and cTnI were increased (P<0.01), the percentage of myocardial infarction area was increased (P<0.01), myocardial fiber was broken and interstitial edema was serious, the positive expression and mRNA expression of GABAA receptor in fastigial nucleus were increased (P<0.01). Compared with the model group, in the EA group, ST segment displacement and LF/HF ratio were decreased (P<0.01), HRV frequency domain analysis showed reduced sympathetic nerve excitability, the serum levels of NE, CK-MB and cTnI were decreased (P<0.01), the percentage of myocardial infarction area was decreased (P<0.01), myocardial fiber breakage and interstitial edema were lightened, the positive expression and mRNA expression of GABAA receptor in fastigial nucleus were decreased (P<0.01). Compared with the EA group, in the agonist group and the agonist+EA group, ST segment displacement and LF/HF ratio were increased (P<0.01), HRV frequency domain analysis showed enhanced sympathetic nerve excitability, the serum levels of NE, CK-MB and cTnI were increased (P<0.01), the percentage of myocardial infarction area was increased (P<0.01), myocardial fiber breakage and interstitial edema were aggravated, the positive expression and mRNA expression of GABAA receptor in fastigial nucleus were increased (P<0.01).@*CONCLUSION@#EA pretreatment can improve the myocardial injury in MIRI rats, and its mechanism may be related to the inhibition of GABAA receptor expression in fastigial nucleus, thereby down-regulating the excitability of sympathetic nerve.

Effect of Tongdu Tiaoshen electroacupuncture pretreatment on PPARγ-mediated pyroptosis of cerebral cortex in rats with cerebral ischemia reperfusion injury / 中国针灸

OBJECTIVE@#To observe the effect of Tongdu Tiaoshen (promoting the circulation of the governor vessel and regulating the spirit) electroacupuncture (EA) pretreatment on pyroptosis mediated by peroxisome proliferators-activated receptor γ (PPARγ) of the cerebral cortex in rats with cerebral ischemia reperfusion injury (CIRI) and explore the potential mechanism of EA for the prevention and treatment of CIRI.@*METHODS@#A total of 110 clean-grade male SD rats were randomly divided into a sham-operation group, a model group, an EA group, an EA + inhibitor group and an agonist group, 22 rats in each group. In the EA group, before modeling, EA was applied to "Baihui" (GV 20), "Fengfu" (GV 16) and "Dazhui" (GV 14), with disperse-dense wave, 2 Hz/5 Hz in frequency, 1 to 2 mA in intensity, lasting 20 min; once a day, consecutively for 7 days. On the base of the intervention as the EA group, on the day 7, the intraperitoneal injection with the PPARγ inhibitor, GW9662 (10 mg/kg) was delivered in the EA + inhibitor group. In the agonist group, on the day 7, the PPARγ agonist, pioglitazone hydrochloride (10 mg/kg) was injected intraperitoneally. At the end of intervention, except the sham-operation group, the modified thread embolization method was adopted to establish the right CIRI model in the rats of the other groups. Using the score of the modified neurological severity score (mNSS), the neurological defect condition of rats was evaluated. TTC staining was adopted to detect the relative cerebral infarction volume of rat, TUNEL staining was used to detect apoptosis of cerebral cortical nerve cells and the transmission electron microscope was used to observe pyroptosis of cerebral cortical neural cells. The positive expression of PPARγ and nucleotide-binding to oligomerization domain-like receptor protein 3 (NLRP3) in the cerebral cortex was detected with the immunofluorescence staining. The protein expression of PPARγ, NLRP3, cysteinyl aspartate specific protease-1 (caspase-1), gasdermin D (GSDMD) and GSDMD-N terminal (GSDMD-N) in the cerebral cortex was detected with Western blot. Using the quantitative real-time fluorescence-PCR, the mRNA expression of PPARγ, NLRP3, caspase-1 and GSDMD of the cerebral cortex was detected. The contents of interleukin (IL)-1β and IL-18 in the cerebral cortex of rats were determined by ELISA.@*RESULTS@#Compared with the sham-operation group, the mNSS, the relative cerebral infarction volume and the TUNEL positive cells rate were increased (P<0.01), pyroptosis was severe, the protein and mRNA expression levels of PPARγ, NLRP3, caspase-1 and GSDMD were elevated (P<0.01); and the protein expression of GSDMD-N and contents of IL-1β and IL-18 were increased (P<0.01) in the model group. When compared with the model group, the mNSS, the relative cerebral infarction volume and the TUNEL positive cells rate were decreased (P<0.01), pyroptosis was alleviated, the protein and mRNA expression levels of PPARγ were increased (P<0.01), the protein and mRNA expression levels of NLRP3, caspase-1 and GSDMD were decreased (P<0.01), the protein expression of GSDMD-N was reduced (P<0.01); and the contents of IL-1β and IL-18 were lower (P<0.01) in the EA group and the agonist group; while, in the EA + inhibitor group, the protein expression of PPARγ was increased (P<0.01), the protein and mRNA expression levels of NLRP3 and GSDMD were decreased (P<0.01, P<0.05), the mRNA expression of caspase-1 was reduced (P<0.01); and the contents of IL-1β and IL-18 were lower (P<0.01). When compared with the EA + inhibitor group, the mNSS, the relative cerebral infarction volume and the TUNEL positive cells rate were decreased (P<0.05, P<0.01), pyroptosis was alleviated, the protein and mRNA expression levels of PPARγ were increased (P<0.01), the protein and mRNA expression levels of NLRP3, caspase-1 and GSDMD were decreased (P<0.01), the protein expression of GSDMD-N was reduced (P<0.01); and the contents of IL-1β and IL-18 were declined (P<0.01) in the EA group. Compared with the agonist group, in the EA group, the relative cerebral infarction volume and the TUNEL positive cells rate were increased (P<0.05, P<0.01), the mRNA expression of PPARγ was decreased (P<0.01) and the protein expression of GSDMD-N was elevated (P<0.05); and the contents of IL-1β and IL-18 were higher (P<0.01).@*CONCLUSION@#Tongdu Tiaoshen EA pretreatment can attenuate the neurological impairment in the rats with CIRI, and the underlying mechanism is related to the up-regulation of PPARγ inducing the inhibition of NLRP3 in the cerebral cortex of rats so that pyroptosis is affected.

Roles of alternative splicing in infectious diseases: from hosts, pathogens to their interactions / 中华医学杂志(英文版)

Alternative splicing (AS) is an evolutionarily conserved mechanism that removes introns and ligates exons to generate mature messenger RNAs (mRNAs), extremely improving the richness of transcriptome and proteome. Both mammal hosts and pathogens require AS to maintain their life activities, and inherent physiological heterogeneity between mammals and pathogens makes them adopt different ways to perform AS. Mammals and fungi conduct a two-step transesterification reaction by spliceosomes to splice each individual mRNA (named cis -splicing). Parasites also use spliceosomes to splice, but this splicing can occur among different mRNAs (named trans -splicing). Bacteria and viruses directly hijack the host's splicing machinery to accomplish this process. Infection-related changes are reflected in the spliceosome behaviors and the characteristics of various splicing regulators (abundance, modification, distribution, movement speed, and conformation), which further radiate to alterations in the global splicing profiles. Genes with splicing changes are enriched in immune-, growth-, or metabolism-related pathways, highlighting approaches through which hosts crosstalk with pathogens. Based on these infection-specific regulators or AS events, several targeted agents have been developed to fight against pathogens. Here, we summarized recent findings in the field of infection-related splicing, including splicing mechanisms of pathogens and hosts, splicing regulation and aberrant AS events, as well as emerging targeted drugs. We aimed to systemically decode host-pathogen interactions from a perspective of splicing. We further discussed the current strategies of drug development, detection methods, analysis algorithms, and database construction, facilitating the annotation of infection-related splicing and the integration of AS with disease phenotype.

Promotion effect of FGF23 on osteopenia in congenital scoliosis through FGFr3/TNAP/OPN pathway / 中华医学杂志(英文版)

BACKGROUND@#Congenital scoliosis (CS) is a complex spinal malformation of unknown etiology with abnormal bone metabolism. Fibroblast growth factor 23 (FGF23), secreted by osteoblasts and osteocytes, can inhibit bone formation and mineralization. This research aims to investigate the relationship between CS and FGF23.@*METHODS@#We collected peripheral blood from two pairs of identical twins for methylation sequencing of the target region. FGF23 mRNA levels in the peripheral blood of CS patients and age-matched controls were measured. Receiver operator characteristic (ROC) curve analyses were conducted to evaluate the specificity and sensitivity of FGF23. The expression levels of FGF23 and its downstream factors fibroblast growth factor receptor 3 (FGFr3)/tissue non-specific alkaline phosphatase (TNAP)/osteopontin (OPN) in primary osteoblasts from CS patients (CS-Ob) and controls (CT-Ob) were detected. In addition, the osteogenic abilities of FGF23-knockdown or FGF23-overexpressing Ob were examined.@*RESULTS@#DNA methylation of the FGF23 gene in CS patients was decreased compared to that of their identical twins, accompanied by increased mRNA levels. CS patients had increased peripheral blood FGF23 mRNA levels and decreased computed tomography (CT) values compared with controls. The FGF23 mRNA levels were negatively correlated with the CT value of the spine, and ROCs of FGF23 mRNA levels showed high sensitivity and specificity for CS. Additionally, significantly increased levels of FGF23, FGFr3, OPN, impaired osteogenic mineralization and lower TNAP levels were observed in CS-Ob. Moreover, FGF23 overexpression in CT-Ob increased FGFr3 and OPN levels and decreased TNAP levels, while FGF23 knockdown induced downregulation of FGFr3 and OPN but upregulation of TNAP in CS-Ob. Mineralization of CS-Ob was rescued after FGF23 knockdown.@*CONCLUSIONS@#Our results suggested increased peripheral blood FGF23 levels, decreased bone mineral density in CS patients, and a good predictive ability of CS by peripheral blood FGF23 levels. FGF23 may contribute to osteopenia in CS patients through FGFr3/TNAP / OPN pathway.

Hypoxia promotes lipopolysaccharide-induced CXCL10 expression in microglia / 生理学报

This study was aimed to investigate the effect of hypoxia on lipopolysaccharide (LPS)-induced CXC-chemokine ligand-10 (CXCL10) expression and the underlying mechanism. C57BL/6J mice were randomly divided into control, hypoxia, LPS, and hypoxia combined with LPS groups. The LPS group was intraperitoneally injected with 0.5 mg/kg LPS, and the hypoxia group was placed in a hypobaric hypoxia chamber (simulated altitude of 6 000 m). The serum and hippocampal tissue samples were collected after 6 h of the treatment. The levels of CXCL10 in the serum and hippocampal tissue of mice were detected by ELISA. The microglia cell line BV2 and primary microglia were stimulated with hypoxia (1% O2) and/or LPS (100 ng/mL) for 6 h. The mRNA expression level of CXCL10 and its content in culture supernatant were detected by real-time quantitative PCR and ELISA, respectively. The phosphorylation levels of nuclear factor κB (NF-κB) signaling pathway-related proteins, p65 and IκBα, were detected by Western blot. Moreover, after NF-κB signaling pathway being blocked with a small molecular compound, PDTC, CXCL10 mRNA expression level was detected in the BV2 cells. The results showed that in the LPS-induced mouse inflammatory model, hypoxia treatment could promote LPS-induced up-regulation of CXCL10 in both serum and hippocampus. Compared with the cells treated with LPS alone, the expression of CXCL10 mRNA and the content of CXCL10 in the culture supernatant of BV2 cells treated with hypoxia combined with LPS were significantly increased. The CXCL10 mRNA level of primary microglial cells treated with hypoxia combined with LPS was significantly up-regulated. Compared with the cells treated with hypoxia or LPS alone, the phosphorylation levels of p65 and IκBα in the BV2 cells treated with hypoxia combined with LPS were significantly increased. PDTC blocked the induction of CXCL10 gene expression by LPS in the BV2 cells. These results suggest that hypoxia promotes LPS-induced expression of CXCL10 in both animal and cell models, and NF-κB signaling pathway plays an important role in this process.

Inhibition of glutaminolysis alleviates myocardial fibrosis induced by angiotensin II / 生理学报

The present study was aimed to investigate the role and mechanism of glutaminolysis of cardiac fibroblasts (CFs) in hypertension-induced myocardial fibrosis. C57BL/6J mice were administered with a chronic infusion of angiotensin II (Ang II, 1.6 mg/kg per d) with a micro-osmotic pump to induce myocardial fibrosis. Masson staining was used to evaluate myocardial fibrosis. The mice were intraperitoneally injected with BPTES (12.5 mg/kg), a glutaminase 1 (GLS1)-specific inhibitor, to inhibit glutaminolysis simultaneously. Immunohistochemistry and Western blot were used to detect protein expression levels of GLS1, Collagen I and Collagen III in cardiac tissue. Neonatal Sprague-Dawley (SD) rat CFs were treated with 4 mmol/L glutamine (Gln) or BPTES (5 μmol/L) with or without Ang II (0.4 μmol/L) stimulation. The CFs were also treated with 2 mmol/L α-ketoglutarate (α-KG) under the stimulation of Ang II and BPTES. Wound healing test and CCK-8 were used to detect CFs migration and proliferation respectively. RT-qPCR and Western blot were used to detect mRNA and protein expression levels of GLS1, Collagen I and Collagen III. The results showed that blood pressure, heart weight and myocardial fibrosis were increased in Ang II-treated mice, and GLS1 expression in cardiac tissue was also significantly up-regulated. Gln significantly promoted the proliferation, migration, mRNA and protein expression of GLS1, Collagen I and Collagen III in the CFs with or without Ang II stimulation, whereas BPTES significantly decreased the above indices in the CFs. α-KG supplementation reversed the inhibitory effect of BPTES on the CFs under Ang II stimulation. Furthermore, in vivo intraperitoneal injection of BPTES alleviated cardiac fibrosis of Ang II-treated mice. In conclusion, glutaminolysis plays an important role in the process of cardiac fibrosis induced by Ang II. Targeted inhibition of glutaminolysis may be a new strategy for the treatment of myocardial fibrosis.

p53 regulates primordial follicle activation through the mTOR signaling pathway / 生理学报

This paper aimed to investigate the role and potential mechanism of p53 on primordial follicle activation. Firstly, the p53 mRNA expression in the ovary of neonatal mice at 3, 5, 7 and 9 days post-partum (dpp) and the subcellular localization of p53 were detected to confirm the expression pattern of p53. Secondly, 2 dpp and 3 dpp ovaries were cultured with p53 inhibitor Pifithrin-μ (PFT-μ, 5 μmol/L) or equal volume of dimethyl sulfoxide for 3 days. The function of p53 in primordial follicle activation was determined by hematoxylin staining and whole ovary follicle counting. The proliferation of cell was detected by immunohistochemistry. The relative mRNA levels and protein levels of the key molecules involved in the classical pathways associated with the growing follicles were examined by immunofluorescence staining, Western blot and real-time PCR, respectively. Finally, rapamycin (RAP) was used to intervene the mTOR signaling pathway, and ovaries were divided into four groups: Control, RAP (1 μmol/L), PFT-μ (5 μmol/L), PFT-μ (5 μmol/L) + RAP (1 μmol/L) groups. The number of follicles in each group was determined by hematoxylin staining and whole ovary follicle counting. The results showed that the expression of p53 mRNA was decreased with the activation of primordial follicles in physiological condition. p53 was expressed in granulosa cells and oocyte cytoplasm of the primordial follicles and growing follicles, and the expression of p53 in the primordial follicles was higher than that in the growing follicles. Inhibition of p53 promoted follicle activation and reduced the primordial follicle reserve. Inhibition of p53 promoted the proliferation of the granulosa cells and oocytes. The mRNA and protein expression levels of key molecules in the PI3K/AKT signaling pathway including AKT, PTEN, and FOXO3a were not significantly changed after PFT-μ treatment, while the expression of RPS6/p-RPS6, the downstream effectors of the mTOR signaling pathway, was upregulated. Inhibition of both p53 and mTOR blocked p53 inhibition-induced primordial follicle activation. Collectively, these findings suggest that p53 may inhibit primordial follicle activation through the mTOR signaling pathway to maintain the primordial follicle reserve.

Angiotensin converting enzyme 2 alleviates infectious bronchitis virus-induced cellular inflammation by suppressing IL-6/JAK2/STAT3 signaling pathway / 生物工程学报

The goal of this study was to investigate the regulatory effect of angiotensin converting enzyme 2 (ACE2) on cellular inflammation caused by avian infectious bronchitis virus (IBV) and the underlying mechanism of such effect. Vero and DF-1 cells were used as test target to be exposed to recombinant IBV virus (IBV-3ab-Luc). Four different groups were tested: the control group, the infection group[IBV-3ab-Luc, MOI (multiplicity of infection)=1], the ACE2 overexpression group[IBV-3ab Luc+pcDNA3.1(+)-ACE2], and the ACE2-depleted group (IBV-3ab-Luc+siRNA-ACE2). After the cells in the infection group started to show cytopathic indicators, the overall protein and RNA in cell of each group were extracted. real-time quantitative polymerase chain reaction (RT-qPCR) was used to determine the mRNA expression level of the IBV nucleoprotein (IBV-N), glycoprotein 130 (gp130) and cellular interleukin-6 (IL-6). Enzyme linked immunosorbent assay (ELISA) was used to determine the level of IL-6 in cell supernatant. Western blotting was performed to determine the level of ACE2 phosphorylation of janus kinase 2 (JAK2) and signal transducer and activator of transcription 3 (STAT3). We found that ACE2 was successfully overexpressed and depleted in both Vero and DF-1 cells. Secondly, cytopathic indicators were observed in infected Vero cells including rounding, detaching, clumping, and formation of syncytia. These indicators were alleviated in ACE2 overexpression group but exacerbated when ACE2 was depleted. Thirdly, in the infection group, capering with the control group, the expression level of IBV-N, gp130, IL-6 mRNA and increased significantly (P < 0.05), the IL-6 level was significant or extremely significant elevated in cell supernatant (P < 0.05 or P < 0.01); the expression of ACE2 decreased significantly (P < 0.05); protein phosphorylation level of JAK2 and STAT3 increased significantly (P < 0.05). Fourthly, comparing with the infected group, the level of IBV-N mRNA expression in the ACE2 overexpression group had no notable change (P > 0.05), but the expression of gp130 mRNA, IL-6 level and expression of mRNA were elevated (P < 0.05) and the protein phosphorylation level of JAK2 and STAT3 decreased significantly (P < 0.05). In the ACE2-depleted group, there was no notable change in IBV-N (P > 0.05), but the IL-6 level and expression of mRNA increased significantly (P < 0.05) and the phosphorylation level of JAK2 and STAT3 protein decreased slightly (P > 0.05). The results demonstrated for the first time that ACE2 did not affect the replication of IBV in DF-1 cell, but it did contribute to the prevention of the activation of the IL-6/JAK2/STAT3 signaling pathway, resulting in an alleviation of IBV-induced cellular inflammation in Vero and DF-1 cells.

Anemoside B4 regulates fatty acid metabolism reprogramming in mice with colitis-associated cancer / 中国中药杂志

The study aimed to investigate the effect of anemoside B4(B4) on fatty acid metabolism in mice with colitis-associated cancer(CAC). The CAC model was established by azoxymethane(AOM)/dextran sodium sulfate(DSS) in mice. Mice were randomly divided into a normal group, a model group, and low-, medium-, and high-dose anemoside B4 groups. After the experiment, the length of the mouse colon and the size of the tumor were measured, and the pathological alterations in the mouse colon were observed using hematoxylin-eosin(HE) staining. The slices of the colon tumor were obtained for spatial metabolome analysis to analyze the distribution of fatty acid metabolism-related substances in the tumor. The mRNA levels of SREBP-1, FAS, ACCα, SCD-1, PPARα, ACOX, UCP-2, and CPT-1 were determined by real-time quantitative PCR(RT-qPCR). The results revealed that the model group showed decreased body weight(P<0.05) and colon length(P<0.001), increased number of tumors, and increased pathological score(P<0.01). Spatial metabolome analysis revealed that the content of fatty acids and their derivatives, carnitine, and phospholipid in the colon tumor was increased. RT-qPCR results indicated that fatty acid de novo synthesis and β-oxidation-related genes, such as SREBP-1, FASN, ACCα, SCD-1, ACOX, UCP-2, and CPT-1 mRNA expression levels increased considerably(P<0.05, P<0.001). After anemoside B4 administration, the colon length increased(P<0.01), and the number of tumors decreased in the high-dose anemoside B4 group(P<0.05). Additionally, spatial metabolome analysis showed that anemoside B4 could decrease the content of fatty acids and their derivatives, carnitine, and phospholipids in colon tumors. Meanwhile, anemoside B4 could also down-regulate the expression of FASN, ACCα, SCD-1, PPARα, ACOX, UCP-2, and CPT-1 in the colon(P<0.05, P<0.01, P<0.001). The findings of this study show that anemoside B4 may inhibit CAC via regulating fatty acid metabolism reprogramming.

Acteoside promotes autophagy and apoptosis of hepatoma cells by regulating JNK signaling pathway / 中国中药杂志

This study explored the molecular mechanism of acteoside against hepatoma 22(H22) tumor in mice through c-Jun N-terminal kinase(JNK) signaling pathway. H22 cells were subcutaneously inoculated in 50 male BALB/c mice, and then the model mice were classified into model group, low-dose, medium-dose, and high-dose acteoside groups, and cisplatin group. The administration lasted 2 weeks for each group(5 consecutive days/week). The general conditions of mice in each group, such as mental status, diet intake, water intake, activity, and fur were observed. The body weight, tumor volume, tumor weight, and tumor-inhibiting rate were compared before and after administration. Morphological changes of liver cancer tissues were observed based on hematoxylin and eosin(HE) staining, and the expression of phosphorylated(p)-JNK, JNK, B-cell lymphoma-2(Bcl-2), Beclin-1, and light chain 3(LC3) in each tissue was detected by immunohistochemistry and Western blot. qRT-PCR was performed to detect the mRNA expression of JNK, Bcl-2, Beclin-1, and LC3. The general conditions of mice in model and low-dose acteoside groups were poor, while the general conditions of mice in the remaining three groups were improved. The body weight of mice in medium-dose acteoside group, high-dose acteoside group, and cisplatin group was smaller than that in model group(P<0.01). The tumor volume in model group was insignificantly different from that in low-dose acteoside group, and the volume in cisplatin group showed no significant difference from that in high-dose acteoside group. Tumor volume and weight in medium-dose and high-dose acteoside groups and cisplatin group were lower than those in the model group(P<0.001). The tumor-inhibiting rates were 10.72%, 40.32%, 53.79%, and 56.44% in the low-dose, medium-dose, and high-dose acteoside groups and cisplatin group, respectively. HE staining showed gradual decrease in the count of hepatoma cells and increasing sign of cell necrosis in the acteoside and cisplatin groups, and the necrosis was particularly obvious in the high-dose acteoside group and cisplatin group. Immunohistochemical results suggested that the expression of Beclin-1, LC3, p-JNK, and JNK was up-regulated in acteoside and cisplatin groups(P<0.05). The results of immunohistochemistry, Western blot, and qRT-PCR indicated that the expression of Bcl-2 was down-regulated in the medium-dose and high-dose acteoside groups and cisplatin group(P<0.01). Western blot showed that the expression of Beclin-1, LC3, and p-JNK was up-regulated in acteoside and cisplatin groups(P<0.01), and there was no difference in the expression of JNK among groups. qRT-PCR results showed that the levels of Beclin-1 and LC3 mRNA were up-regulated in the acteoside and cisplatin groups(P<0.05), and the level of JNK mRNA was up-regulated in medium-dose and high-dose acteoside groups and cisplatin group(P<0.001). Acteoside promotes apoptosis and autophagy of H22 cells in mice hepatoma cells by up-regulating the JNK signaling pathway, thus inhibiting tumor growth.