Redox metabolism maintains the leukemogenic capacity and drug resistance of AML cells.

Rewiring of redox metabolism has a profound impact on tumor development, but how the cellular heterogeneity of redox balance affects leukemogenesis remains unknown. To precisely characterize the dynamic change in redox metabolism in vivo, we developed a bright genetically encoded biosensor for H2O2 (named HyPerion) and tracked the redox state of leukemic cells in situ in a transgenic sensor mouse. A H2O2-low (HyPerion-low) subset of acute myeloid leukemia (AML) cells was enriched with leukemia-initiating cells, which were endowed with high colony-forming ability, potent drug resistance, endosteal rather than vascular localization, and short survival. Significantly high expression of malic enzymes, including ME1/3, accounted for nicotinamide adenine dinucleotide phosphate (NADPH) production and the subsequent low abundance of H2O2. Deletion of malic enzymes decreased the population size of leukemia-initiating cells and impaired their leukemogenic capacity and drug resistance. In summary, by establishing an in vivo redox monitoring tool at single-cell resolution, this work reveals a critical role of redox metabolism in leukemogenesis and a potential therapeutic target.

Saponins from Panax japonicus improve neuronal mitochondrial injury of aging rats.

CONTEXT: Panax japonicus is the dried rhizome of Panax japonicus C.A. Mey. (Araliaceae). Saponins from Panax japonicus (SPJ) exhibit anti-oxidative and anti-aging effects. OBJECTIVE: We evaluated the neuroprotective effects of SPJ on aging rats. MATERIALS AND METHODS: Sprague-Dawley rats (18-months-old) were randomly divided into aging and SPJ groups (n = 8). Five-month-old rats were taken as the adult control (n = 8). The rats were fed a normal chow diet or the SPJ-containing diet (10 or 30 mg/kg) for 4 months. An in vitro model was established by d-galactose (d-Gal) in the SH-SY5Y cell line and pretreated with SPJ (25 and 50 µg/mL). The neuroprotection of SPJ was evaluated via Nissl staining, flow cytometry, transmission electron microscopy and Western blotting in vivo and in vitro. RESULTS: SPJ improved the neuronal degeneration and mitochondrial morphology that are associated with aging. Meanwhile, SPJ up-regulated the protein levels of mitofusin 2 (Mfn2) and optic atrophy 1 (Opa1) and down-regulated the protein level of dynamin-like protein 1 (Drp1) in the hippocampus of aging rats (p < 0.05 or p < 0.01 vs. 22 M). The in vitro studies also demonstrated that SPJ attenuated d-Gal-induced cell senescence concomitant with the improvement in mitochondrial function; SPJ, also up-regulated the Mfn2 and Opa1 protein levels, whereas the Drp1 protein level (p < 0.05 or p < 0.01 vs. d-Gal group) was down-regulated. DISCUSSION AND CONCLUSIONS: Further research on the elderly population will contribute to the development and utilization of SPJ for the treatment of neurodegenerative disorders.

A single-component light sensor system allows highly tunable and direct activation of gene expression in bacterial cells.

Light-regulated modules offer unprecedented new ways to control cellular behaviour with precise spatial and temporal resolution. Among a variety of bacterial light-switchable gene expression systems, single-component systems consisting of single transcription factors would be more useful due to the advantages of speed, simplicity, and versatility. In the present study, we developed a single-component light-activated bacterial gene expression system (eLightOn) based on a novel LOV domain from Rhodobacter sphaeroides (RsLOV). The eLightOn system showed significant improvements over the existing single-component bacterial light-activated expression systems, with benefits including a high ON/OFF ratio of >500-fold, a high activation level, fast activation kinetics, and/or good adaptability. Additionally, the induction characteristics, including regulatory windows, activation kinetics and light sensitivities, were highly tunable by altering the expression level of LexRO. We demonstrated the usefulness of the eLightOn system in regulating cell division and swimming by controlling the expression of the FtsZ and CheZ genes, respectively, as well as constructing synthetic Boolean logic gates using light and arabinose as the two inputs. Taken together, our data indicate that the eLightOn system is a robust and highly tunable tool for quantitative and spatiotemporal control of bacterial gene expression.

Strong valley splitting ind0two-dimensional SnO induced by magnetic proximity effect.

Strong magnetic interfacial coupling in van der Waals heterostructures is important for designing novel electronic devices. Besides the most studied transition metal dichalcogenides (TMDCs) materials, we demonstrate that the valley splitting can be activated in two-dimensional tetragonald0metal oxide, SnO, via the magnetic proximity effect by EuBrO. In SnO/EuBrO, the valley splitting of SnO can reach ∼46 meV, which is comparable to many TMDCs and equivalent to an external magnetic field of 800 T. In addition, the valley splitting can be further enhanced by adjusting interlayer distance and applying uniaxial strains. A design principle of new spintronic device based on this unique electronic structure of SnO/EuBrO has been proposed. Our findings indicate that SnO is a promising material for future valleytronics applications.

Icariin improves brain function decline in aging rats by enhancing neuronal autophagy through the AMPK/mTOR/ULK1 pathway.

CONTEXT: Icariin (ICA) is the main active ingredient of Epimedium brevicornu Maxim (Berberidaceae), which is used in the immune, reproductive, neuroendocrine systems, and anti-aging. OBJECTIVE: To evaluate the effect of ICA on natural aging rat. MATERIALS AND METHODS: 16-month-old Sprague-Dawley (SD) rats were randomly divided into aging, low and high-dose ICA groups (n = 8); 6-month-old rats were taken as the adult control (n = 8). Rats were fed regular feed (aging and adult control) or feed containing ICA (ICA 2 and 6 mg/kg group) for 4 months. HE and Nissl staining were used to assess pathological changes. Western blot was used to test the expression of autophagy (LC3B, p62, Atg5, Beclin1) and p-AMPK, p-mTOR and p-ULK1 (ser 757). Immunofluorescence was used to detect the co-localization of LC3 and neurons. RESULTS: ICA improved neuronal degeneration associated with aging and increased the staining of Nissl bodies. Western blot showed that ICA up-regulated autophagy-related proteins LC3B (595%), Beclin1 (73.5%), p-AMPK (464%) protein (p < 0.05 vs. 20 M) in the cortex and hippocampus of aging rats, down-regulated the expression of p62 (56.9%), p-mTOR (53%) and p-ULK1 (ser 757) (65.4%) protein (p < 0.05 vs. 20 M). Immunofluorescence showed that the fluorescence intensity of LC3 decreased in the aging rat brain, but increased and mainly co-localized with neurons after ICA intervention. CONCLUSIONS: Further research needs to verify the expression changes of AMPK/mTOR/ULK1 and the improvement effect of ICA in elderly. These results will further accelerate the applications of ICA and the treatment for senescence.

Saponins from Panax japonicus attenuate cognitive impairment in ageing rats through regulating microglial polarisation and autophagy.

CONTEXT: Panax japonicus is the dried rhizome of Panax japonicus C.A. Mey. (Araliaceae). Saponins from Panax japonicus (SPJ) exhibit anti-inflammatory and antioxidative effects. OBJECTIVE: To explore the neuroprotective effect of SPJ on natural ageing of rat. MATERIALS AND METHODS: Sprague-Dawley (SD) rats 18-month-old were divided into ageing control, ageing treated with SPJ 10 or 30 mg/kg (n = 8). Five-month-old rats were taken as the adult control (n = 8). Rats were fed regular feed or feed containing SPJ for 4 months. Cognitive level was evaluated by Morris water maze (MWM) test. The mechanisms of SPJ's neuroprotection were evaluated by transmission electron microscope, western blot analysis, and immunofluorescence in vivo and in vitro. RESULTS: SPJ attenuated ageing-induced cognitive impairment as indicated by elevated number of times crossing the target platform (from 1.63 to 3.5) and longer time spent in the target platform quadrant (from 1.33 to 1.98). Meanwhile, SPJ improved the morphology of microglia and synapse, and activated M2 microglia polarisation including increased hippocampus levels of CD206 (from 0.98 to 1.47) and YM-1 (from 0.67 to 1.1), and enhanced autophagy-related proteins LC3B (from 0.48 to 0.82), Beclin1 (from 0.32 to 0.51), Atg5 (from 0.22 to 0.89) whereas decreased p62 level (from 0.71 to 0.45) of ageing rats. In vitro study also showed that SPJ regulated the microglial polarisation and autophagy. DISCUSSION AND CONCLUSIONS: SPJ improved cognitive deficits of ageing rats through attenuating microglial inflammation and enhancing microglial autophagy, which could be used to treat neurodegenerative disorders.

[Effect of total saponins from Panax japonicus on non-alcoholic steatohepatitis by regulating autophagy].

Non-alcoholic steatohepatitis(NASH) was induced by high-sugar and high-fat diet in mice to investigate the intervention effect of total saponins from Panax japonicus(TSPJ) and explore its possible mechanism. Mice were fed with high-sugar and high-fat diet to establish NASH model, and intervened with different doses of TSPJ(15, 45 mg·kg~(-1)). The animals were fed for 26 weeks. The histomorphology and pathological changes of liver tissues were observed by HE staining. The transcriptional expression levels of miR-199 a-5 p, autophagy related gene 5(ATG5) and inflammatory cytokines interleukin-6(IL-6), interleukin-1ß(IL-1ß) and tumor necrosis factor α(TNF-α) in mouse liver were measured by quantitative Real-time polymerase chain reaction(qRT-PCR). Western blot was used to detect the expression of autophagy-related proteins ATG5, P62/SQSTM1(P62), and microtubule-associated protein light chain 3(LC3)-I/Ⅱ proteins in mouse liver. The expression of P62 protein was detected by immunofluorescence staining. In order to verify the targeting regulation relationship between miR-199 a-5 p and ATG5, miR mimic/inhibitor NC and miR-199 a-5 p mimic/inhibitor were transfected into Hepa 1-6 cells, and the expression of ATG5 mRNA and protein was detected. pMIR-reportor ATG5-3'UTR luciferase reporter gene plasmid was constructed and co-transfected with miR mimic/inhibitor NC and miR-199 a-5 p mimic/inhibitor into Hepa 1-6 cells to detect luciferase activity. In vivo, HE staining in the model group showed typical fatty degeneration and inflammatory infiltration, with increased expression of miR-199 a-5 p and decreased expression of ATG5 mRNA and protein. The expression of autophagy-associated protein P62 increased significantly, the ratio of LC3Ⅱ/Ⅰ decreased, and the transcriptional expression of inflammatory factors increased significantly. After the intervention by TSPJ, the pathological performance of liver tissue was significantly improved, the expression of miR-199 a-5 p decreased and the expression of ATG5 mRNA and protein increased, the expression of autophagy-associated protein P62 decreased significantly, the ratio of LC3Ⅱ/Ⅰ increased, and the transcriptional expression of inflammatory cytokines IL-6, IL-1ß and TNF-α decreased significantly. In vitro, it was found that the expression of ATG5 mRNA and protein and luciferase activity decreased significantly in miR-199 a-5 p overexpression cells, while after inhibition of miR-199 a-5 p expression, the expression level of ATG5 mRNA and protein and luciferase activity increased. The results showed that TSPJ can improve NASH in mice fed with high-sugar and high-fat diet, and its mechanism may be related to the regulation of miR-199 a-5 p/ATG5 signal pathway, the regulation of autophagy activity and the improvement of inflammatory response of NASH.

A 2D Rashba electron gas with large spin splitting in Janus structures of SnPbO2.

For realizing two-dimensional (2D) spintronic devices, controllable manipulation of different spin channels is needed. In order to achieve this goal, the materials should have well-separated spin energies induced by spin-orbit coupling (SOC). However, a negligible SOC effect in 2D SnO limits its application in this prospect. Herein, we demonstrate that the stable Janus structure of SnPbO2 can exhibit large spin splitting (67 meV) at the valence band maximum (VBM) due to the breakdown of inverse symmetry. This spin splitting is larger than the thermal fluctuation at room temperature as well as some other 2D systems. In addition, the splitting can be further enhanced by strains (82 meV) or hole doping (147 meV). More importantly, there are no other electronic states except for the Rashba states at the VBM, which make it ideal for practical applications. Our study provides alternative ways to regulate the electronic structure of SnO and the feasibility for spin manipulation in this interesting 2D system.

Imaging the Redox States of Live Cells with the Time-Resolved Fluorescence of Genetically Encoded Biosensors.

Redox environments in cells influence many important physiological and pathological processes. In this study, the time-resolved fluorescence of a recently reported thiol redox-sensitive sensor based on vertebrate fluorescent protein UnaG, roUnaG, was studied, along with the application of the time-resolved fluorescence of roUnaG to image the redox states of the mitochondria, cytoplasm, and nucleus in live cells. Time-resolved fluorescence images of roUnaG clearly demonstrated that potent anticancer compound KP372-1 induced extreme oxidative stress. A more stressful redox state observed in activated macrophages further demonstrated the validity of roUnaG with time-resolved fluorescence. For comparison, time-resolved fluorescence images of four other frequently used redox biosensors (roGFP1, HyPer, HyPerRed, and rxRFP) were also captured. The time-resolved fluorescence allows an intrinsically ratiometric measurement for biosensors with one excitation wavelength and provides new opportunities for bioimaging.

Ultrafast internal conversion dynamics of bilirubin bound to UnaG and its N57A mutant.

Fluorescent proteins (FPs) have become fundamental tools for live cell imaging. Most FPs currently used are members of the green fluorescent protein super-family, but new fluorophores such as bilin-FPs are being developed and optimized. In particular, the UnaG FP incorporates bilirubin (BR) as a chromophore, enhancing its fluorescence quantum yield by three orders of magnitude relative to that in solution. To investigate the mechanism of this dramatic enhancement and provide a basis for further engineering of UnaG and other tetrapyrrole-based fluorophores, we performed picosecond fluorescence and femtosecond transient absorption measurements of BR bound to UnaG and its N57A site-directed mutant. The dynamics of wt-UnaG, which has a fluorescence QY of 0.51, are largely homogeneous, showing an excited state relaxation of ∼200 ps, and a 2.2 ns excited-state lifetime decay with a kinetic isotope effect (KIE) of 1.1 for D2O vs. H2O buffer. In contrast, for UnaG N57A (fluorescence QY 0.01) the results show a large spectral inhomogeneity with excited state decay timescales of 47 and 200 ps and a KIE of 1.4. The non-radiative deactivation of the excited state is limited by proton transfer. The loss of direct hydrogen bonds to the endo-vinyl dipyrrinone moiety of BR leads to high flexibility and structural heterogeneity of UnaG N57A, as seen in the X-ray crystal structure.

Protective effect of Wuzi Yanzong recipe on testicular dysfunction through inhibition of germ cell apoptosis in ageing rats via endoplasmic reticulum stress.

It has been demonstrated that excessively activated endoplasmic reticulum stress (ERS) is closely associated with ageing-related diseases and male reproductive dysfunction. Wuzi Yanzong recipe (WZ) is a classical Traditional Chinese Medicine prescription for treatment of male reproductive system diseases. However, it remains unknown whether WZ improves testicular dysfunction with ageing via ERS. In this study, we investigated the protective effects and its mechanism of WZ on testicular dysfunction in ageing rats. The results showed that treatment with WZ for 4 months significantly increased the testicular weight and index, sperm count and viability, and the levels of testosterone and decreased the levels of estradiol. In addition, WZ significantly activated the onset of ERS and prevented germ cell apoptosis by upregulating the expression levels of ERS-responsive proteins GRP78, phospho-PERK, phospho-eIF2α, ATF4, phospho-IRE-1α, XBP1 and ATF6α, and downregulating the expression levels of pro-apoptotic proteins p-JNK, Caspase12 and CHOP in testicular germ cell of ageing rats. Besides, WZ significantly decreased the numbers of TUNEL-positive cells. Taken together, WZ effectively improves ageing-related testicular dysfunction through inhibition of germ cell apoptosis via ERS.

[Triterpene saponins in Panax japonicus and their ~(13)C-NMR spectroscopic characteristics].

Panax japonicus( PJ) is a valuable medicinal plant belonging to the genus Panax of Araliaceae,the recumbent rhizome of which is widely used in clinic therapy,healthcare products and as cosmetic additives with functions of dissipating stasis,reducing swelling,stanching bleeding,and reinforcing deficiency,etc. PJ contains abundant levels of oleanane-and dammarane-type triterpene saponins,which are considered as the material basis for exerting pharmacodynamic action. Based on the previous researches,more than110 triterpene saponins have been reported from PJ. These triterpene saponins were summarized in this review,and could be classified into dammarenediol Ⅱ,protopanaxadiol,protopanaxatiol,ocotillol,oleanolic acid,ursolic acid and miscellaneous subtypes,according to their molecular skeletons in biosynthesis processes. Further more,the structural features of these triterpene saponins in the seven different subtypes,together with their~(13)C-NMR spectroscopic characteristics were described,hoping to provide available information for chemical diversity research of PJ.

[Improved effects of saponins from Panax japonicus on decline of cognitive function in natural aging rats via NLRP3 inflammasome pathway].

The aim of this paper was to investigate the effect of total saponins from Panax japonicus( SPJ) on cognitive decline of natural aging rats and its mechanism. Thirty male SD rats of eighteen month old were randomly divided into three groups: aged group,10 mg·kg~(-1) SPJ-treated group and 30 mg·kg~(-1) SPJ-treated group. The SPJ-treated groups were given SPJ at the dosages of 10 mg·kg~(-1) and 30 mg·kg~(-1),respectively,from the age of 18 to 24 months. Aged group were lavaged the same amount of saline,10 six-month-old rats were used as control group,with 10 rats in each group. The open field test,novel object recognition and Morris water maze were performed to detect the changes of cognitive function in each group. The changes of synaptic transmission of long-term potentiation( LTP) in hippocampal CA1 region were detected by field potential recording. Western blot was used to detect the protein levels of NLRP3,ASC,caspase-1 and the changes of Glu A1,Glu A2,CAMKⅡ,CREB and phosphorylation of CAMKⅡ,CREB in each group.The results showed that SPJ could improve the decline of cognitive function in aging rats,reduce the damage of LTP in the hippocampal CA1 region of aged rats,and decrease the expression of NLRP3,ASC,caspase-1 in aging rats. At the same time,SPJ could enhance the membrane expression of AMPA receptor( Glu A1 and Glu A2),and increase the expression of p-CAMKⅡand p-CREB in aging rats.SPJ could improve cognitive decline of natural aging rats,and its mechanism may be related to regulating NLRP3 inflammasome,thus regulating the membrane expression of AMPA receptor,and enhancing the expression phosphorylation of CAMKⅡ and CREB.

Panax notoginseng saponins attenuate cardiomyocyte apoptosis through mitochondrial pathway in natural aging rats.

Panax notoginseng saponins (PNS) have been widely used in the cardiovascular system for the treatment of cardiovascular diseases and stroke in China. In this study, we investigated the anti-apoptotic effect of PNS on cardiomyocytes in the natural aging rat and explored the potential mechanisms regarding oxidative stress and mitochondrial function signaling pathways. Male Sprague-Dawley rats were randomly divided into five groups: adult control (3-month old), aging control (24-month old), and different doses of PNS-treated aging rat groups (10, 30, 60 mg/kg/day, orally). After treatment of PNS or saline for 6 months, the effects of PNS on the cardiomyocytes were evaluated. Results showed that PNS significantly improved the morphological changes in myocardium, prevented the increase of cardiomyocyte apoptosis in the aging rats, and improved mitochondrial dysfunction associated aging in a dose-dependent manner. PNS also significantly reversed the down-regulation of FoxO3a and Mn-SOD and up-regulated PGC-1α, LC3ß, and Beclin-1 levels. Our data demonstrated that during aging, mitochondrial dysfunction caused an increase of oxidative damage, which played a key role in cardiomyocyte apoptosis. PNS exerted an anti-apoptotic effect via attenuating oxidative damage through oxidative stress- and mitochondrial function-related signaling pathways.

[Effect of saponins extracted from Panax japonicas on inhibiting myocardial fibrosis by TGF-ß1/Smad3 signaling pathway in aging rats].

To investigate the amelioration effect of saponins extracted from Panax japonicas (SPJ) on myocardial fibrosis in natural aging rats and its mechanisms, male SD rats aged 18 months were randomly divided into 3 groups (aging model group, low-dose SPJ group and high-dose SPJ group), with 10 rats in each group. SPJ groups were given SPJ at different doses (10, 60 mg·kg⁻¹·d⁻¹) consecutively for 6 months, meanwhile, aging model group was treated with the equal volume of saline for 6 months until 24 months old. Another 10 rats aged 6 month were used as young control group. The changes of myocardial morphological were observed by haematoxylin-eosin (HE) staining. Masson staining was used to observe the changes of collagen deposition in rat hearts. RT-PCR was used to detect the mRNA expression levels of myofibroblast marker α-SMA, collagen-related protein COL1α2, COL3α1 and matrix metalloproteinase MMP2, MMP9. Western blot was used to test the changes of the protein expressions of TGF-ß1, p-Smad3, IL-1ß and TNF-α in heart tissues. SPJ can effectively improve the arrangement of myocardial fibers, decrease inflammatory infiltration and reduce collagen deposition in aging rats. SPJ can effectively down-regulate the mRNA expression levels of COL1α2, COL3α1, α-SMA, MMP9, MMP2 and inhibit the protein expressions of TGF-ß1, p-Smad3, TNF-α, IL-1ß in the natural aging heart tissues. SPJ can effectively alleviate myocardial fibrosis in natural aging rats, and its mechanisms was related to the inhibition of the protein expressions of TGF-ß1, p-Smad3 and the reduction of myocardial inflammation in rat hearts.

[Protective effect of Wuzi Yanzong recipe on testicular germ cell apoptosis in natural ageing rats through endoplasmic reticulum stress].

To study the protective effects of Wuzi Yanzong recipe on testis germ cell apoptosis in natural ageing rats through endoplasmic reticulum stress (ERS), 16-month-old male SPF grade SD rats were randomly divided into three groups: ageing model group, and low and high-dose Wuzi Yanzong recipe groups (WZ, 1 and 4 g·kg⁻¹), with 10 rats in each group. In addition, 2-month-old SD male rats were used as adult control group. The ageing model group and the adult control group were fed with normal diet for 4 months. WZ groups were given the medicated feed for 4 months. After fasting for 12 hours, the rats were put to death. Then, the testes were immediately collected. The change of testicular tissue morphology was observed by HE staining. The expression levels of ER stress-related proteins GRP78, p-PERK, p-eif2α, ATF4, p-IRE1, XBP1, ATF6 and apoptosis-related proteins CHOP, caspase12 and p-JNK in testes were detected by Western blot. Compared with the ageing model group, Wuzi Yanzong recipe alleviated the morphological changes of testicular tissue. Western blot results showed that Wuzi Yanzong recipe significantly increased the expression levels of endoplasmic reticulum stress-related proteins GRP78, p-PERK, p-eif2α, ATF4, p-IRE1, XBP1, ATF6 and significantly decreased the expression levels of endoplasmic reticulum-induced apoptosis-related proteins CHOP, caspase 12 and p-JNK. In conclusion, Wuzi Yanzong recipe can alleviate the ageing-related apoptosis of testicular germ cells in natural ageing rats by regulating endoplasmic reticulum stress.

[Protective effect of Wuzi Yanzong recipe on testicular DNA oxidative damage in natural ageing rats].

To study the protective effects of Wuzi Yanzong recipe on DNA oxidative damage of testis germ cells in natural ageing rats based on Nrf2/HO-1 signaling pathway and base excision repair (BER). In the study, 16-month-old SPF grade male SD rats were randomly divided into three groups, namely ageing model group, and low and high-dose Wuzi Yanzong recipe groups (WZ, 1, 4 g·kg⁻¹). In addition, 2-month-old SD rats were used as adult control group (10 rats in each group). The ageing model group and the adult control group were fed with normal diet for 4 months. WZ groups were given medicated feed for 4 months. After fasting for 12 hours, the rats were put to death. Then, the testes were immediately removed. The vitality of superoxide dismutase (SOD) and malondialdehyde (MDA) content in testis were detected by xanthine oxidase method and thiobarbituric acid (TBA) method. The levels of Nrf2 and 8-OHdG were detected by immunofluorescence. The protein expression levels of Nrf2, HO-1, NQO1, APE1, OGG1 and XRCC1 were detected by Western blot. Compared with the ageing model group, WZ significantly increased the SOD vitality and decreased MDA content of testis. In addition, immunofluorescence results showed that WZ significantly attenuated testicular DNA oxidative damage and improved antioxidant capacity. Such changes were accompanied by the down-regulation of DNA oxidative damage response protein 8-OHdG levels and the up-regulation of Nrf2 levels. Moreover, Western blot results showed that WZ significantly increased the protein expression levels of Nrf2, HO-1 and NQO1 of the testis germ cells, when compared with ageing model group. In parallel, the protein expression levels of APE1, OGG1 and XRCC1 were significantly decreased. In conclusion, WZ improves ageing-related DNA oxidative damage via Nrf2/HO-1 and BER pathways.

[Protective effect of Wuzi Yanzong recipe on testicular DNA damage and apoptosis in natural ageing rats].

To study the protective effect of Wuzi Yanzong recipe on testicular DNA damage and apoptosis in natural ageing rats, SPF grade 16-month-old SD male rats were randomly divided into three groups: ageing model group, low and high dose Wuzi Yanzong recipe groups (WZ, 1， 4 g·kg⁻¹). In addition, 2-month-old SD rats were used as adult control group (10 rats in each group). The ageing model group and the adult control group were fed with normal diet for 4 months. Wuzi Yanzong groups received medicated feed for 4 months. After fasting for 12 hours, the rats were sacrificed. Then testis tissues were taken and weighed to calculate the testis index. The change of testicular tissue morphology was observed by HE staining. Expression and localization of DNA damage-associated protein ATR were observed by immunofluorescence. The expressions of DNA damage-related proteins γ-H2AX, Chk1, p-p53 and apoptosis-related proteins Bcl-2 and Bax in testes were detected by Western blot. The apoptosis of testis tissue in rats was detected by using TUNEL. The results showed that as compared with the youth control group, the protein expression levels of γ-H2AX, Chk1, p-p53 and Bax were significantly increased while Bcl-2 protein expression level was significantly decreased intestis tissues of ageing model group. Wuzi Yanzong recipe significantly decreased protein expression levels of γ-H2AX, Chk1, p-p53 and Bax and increased Bcl-2 protein expression level as well as Bcl-2/Bax ratio. Immunofluorescence results showed that Wuzi Yanzong recipe could significantly decrease the ageing-induced ATR, increase in testis tissues. TUNEL results showed that Wuzi Yanzong recipe could significantly attenuate the germ cell apoptosis in testicular tissues. All the above results suggest that Wuzi Yanzong recipe could protect the germ cell in testicular tissues of natural ageing rates from DNA damage and apoptosis, and the mechanism may be associated with regulating p53 signaling pathway.

[Protective effects of total saponins of Panax japonicas on HepG2 cell apoptosis induced with palmitic acid].

This study aimed to investigate the molecular mechanism and protective effect of total saponins of Panax japonicas (TSPJ) on HepG2 cells apoptosis induced by palmitic acid (PA).The HepG2 cells were cultured in vitro, and divided into five groups: the control group, the model group, the high-dose group (50 mg·L⁻¹), the middle-dose group (25 mg·L⁻¹) and the low-dose group (12.5 mg·L⁻¹).The cells of the five groups were cultured continuously for 24 hours. The cell viability was measured with MTT. HepG2 cells apoptosis was detected by Hoechest staining and Annexin V-FITC/PI staining. The protein expressions of BCL-2, CHOP and TLR4 were measured with western blotting and flow cytometry analysis. The mRNA expressions of TNF-α, IL-1ß, BCL-2, CHOP and GAPDH were measured with RT-PCR. The results suggested that compared with the control group, the number of HepG2 cells of the model group were reduced significantly (P<0.01), while the number of apoptotic HepG2 cells were increased. Compared with the model group, the number of HepG2 cells of the high-dose group and the middle-dose group were increased significantly (P<0.01), whereas the number of apoptotic HepG2 cells were reduced. Compared with the control group, TNF-α, IL-1ß and CHOP mRNA expressions and CHOP and TLR4 protein expressions in the model group were significantly up-regulated (P<0.01), while BCL-2 protein and mRNA expressions in the model group were significantly decreased (P<0.01). Compared with the model group, TNF-α, IL-1ß and CHOP mRNA expressions and CHOP and TLR4 protein expressions in the high-dose group were significantly decreased (P<0.01), while BCL-2 protein and mRNA expressions in the high-dose group were significantly up-regulated (P<0.01).In conclusion, TSPJ can reduce inflammation and apoptosis induced by palmitic acid, with a certain protective effect on liver cells.

[Protective effects of total saponins of Panax japonicas on non-alcoholic fatty liver disease through regulating expression of miR-199-5p].

To research the effection and probable mechanism for the total saponins of Panax japonicas(TPSJ) in mice on non-alcoholic fatty liver disease. Forty SPF male Kunming mice were randomily divided into four group:control group,NAFLD group, low-dose TPSJ treated group,high-dose TPSJ treated group. High-fatty and high-frutose-diet was applied to eatablish NAFLD model，and TPSJ (100 and 200 mg·kg⁻¹) in feeding were given for the TPSJ groups for 4 weeks. To collect the serum with liver and the ALT and TC of serum were monitored after 4 weeks. The hepatic histopathologic structure was observed by haematoxylin-eosin (HE) staining, RT-PCR and RT-qPCR was applied for the detection of miR-199-5p，VEGFa，HGF，c-Met and protein expression level was detected bv laser confocal microscope.Compared with control group, the level of serum ALT and TC in the model group was higher,the liver of the model group showed that hepatocytes display obvious lipid deposition. Then TPSJ treated showed that markedly improved histopathologic changes, decreased fatty deposition. In the meantime,the expression level of miR-199-5p was significantly decreased, thus the expression of HGF and c-Met were significantly increased. TPSJ play a role of prevention on fatty liver, the machanism maybe by blocking miR-199-5p targeted to c-Met signaling pathways in NAFLD.