Effect of Bushen Huoxue recipe on women with thin endometrial ovulation disorder and a rat model of thin endometrium resulted from kidney deficiency-related blood stasis.

To examine the therapeutic effect of Bushen Huoxue recipe (BHR) on women with thin endometrial ovulation disorder and on a rat model of kidney deficiency-related blood stasis. A total of 60 women with thin endometrial ovulation disorder was enrolled. The primary outcome of the study was the pregnancy rate three menstrual cycles after treatment. The study also examined the changes in the type and thickness of uterine artery, uterine artery pulsatility index (PI) and endometrial resistance index (RI). To establish kidney deficiency-related blood stasis in Sprague Dawley (SD) rats, an intragastric administration of hydroxyurea and a tail vein injection of Dextran were given, following with a flashing of the uterine cavity with 95% anhydrous ethanol. A combined regimen of BHR and estradiol valerate significantly increased the rate of pregnancy in women with thin endometrial ovulation disorder. The treatment was accompanied by a significant increase in endometrial thickness and decreases in uterine artery PI and endometrial RI. In rats, kidney deficiency-related blood stasis caused severe loss in endometrial architecture, thickness, and numbers of gland and blood vessel compared to the healthy SD rats. Treatment with BHR could ameliorate the endometrial damages associated with kidney deficiency-related blood stasis.

Paeoniflorin ameliorates glycemic variability-induced oxidative stress and platelet activation in HUVECs and DM rats.

Glycemic variability (GV) plays an important role in the pathogenesis of vascular complications associated with diabetes mellitus (DM). Paeoniflorin is an effective Chinese traditional medicine with anti-inflammatory and immune-regulatory effects. Previous studies implicated the beneficial effects of paeoniflorin in treatment for diabetic complications, such as type 2 diabetic nephropathy and diabetes with myocardial ischemic injury. Current evidence suggests that oxidative stress and platelet activation, as well as their interaction, are potentially associated with GV and involved in the pathogenesis of diabetes-associated vascular complications. This study aimed to explore the effects of paeoniflorin on oxidative stress and platelet activation, using human umbilical vein endothelial cells (HUVECs) cultured with different glucose concentrations, and streptozotocin-induced diabetic rats fed different glycemic index diets. Paeoniflorin treatment effectively improved the morphology and cell viability of HUVECs under glucose fluctuation. Moreover, the platelet aggregation rate, CD62p expression, and reactive oxygen species (ROS) concentration decreased, while glutathione peroxidase (GSH-px) levels increased in paeoniflorin-treated groups. In conclusion, our study found that paeoniflorin ameliorates oxidative stress and platelet activation induced by glycemic variability both in vivo and in vitro, suggesting a novel potential strategy for treatment of diabetic complications.

A Protective Role of Paeoniflorin in Fluctuant Hyperglycemia-Induced Vascular Endothelial Injuries through Antioxidative and Anti-Inflammatory Effects and Reduction of PKCß1.

Hyperglycemia fluctuation is associated with diabetes mellitus (DM) complications when compared to persistent hyperglycemia. Previous studies have shown that paeoniflorin (PF), through its antiapoptosis, anti-inflammation, and antithrombotic properties, effectively protects against cardiovascular and cerebrovascular disease. However, the mechanism underlying the protection from PF against vascular injuries induced by hyperglycemia fluctuations remains poorly understood. Herein, we investigated the potential protective role of PF on human umbilical vein endothelial cells (HUVECs) subjected to intermittent glucose levels in vitro and in DM rats with fluctuating hyperglycemia in vivo. A remarkable increased apoptosis associated with elevated inflammation, increased oxidative stress, and high protein level of PKCß1 was induced in HUVECs by intermittently changing glucose for 8 days, and PF recovered those detrimental changes. LY333531, a potent PKCß1 inhibitor, and metformin manifested similar effects. Additionally, in DM rats with fluctuating hyperglycemia, PF protected against vascular damage as what has been observed in vitro. Taken together, PF attenuates the vascular injury induced by fluctuant hyperglycemia through oxidative stress inhibition, inflammatory reaction reduction, and PKCß1 protein level repression, suggesting its perspective clinical usage.

[Advances in the molecular genetic studies of acephalic spermatozoa syndrome].

Acephalic spermatozoa syndrome (ASS) is characterized by a predominance of headless spermatozoa with abnormal head-tail junction in the ejaculate, which causes severe male infertility. The pathogenic mechanism of ASS remained unclarified for a long time until recent identification of the four ASS-associated genes SUN5, PMFBP1, TSGA10, and BRDT and their mutations due to the development of high-throughput sequencing technology. This review summarizes the advances in the genetic studies of ASS, focusing on its pathogenic molecular mechanisms, which provide an important basis for the molecular diagnosis of the disease as well as for assisted reproductive technology.

Effect of tetramethylpyrazine and hyperlipidemia on hepcidin homeostasis in mice.

Iron homeostasis is strictly regulated in mammals, and disordered iron metabolism is recognized as a risk factor for various diseases, including cardiovascular disease. The hepcidin­ferroportin axis is the key signaling mechanism that controls systemic iron homeostasis. Increased serum hepcidin is associated with multiple types of cancer and atherosclerosis (AS), and therapeutics that decrease hepcidin levels have been proposed to treat these diseases. However, the effects of abnormal circulating hepcidin on hyperlipidemia remain unexploited. The natural compound tetramethylpyrazine (TMP) has been reported to have therapeutic effects on cardiovascular diseases, whereas the mechanisms involved remain incompletely understood. Thus, the effects of TMP on the expression of hepcidin in hyperlipidemic mice were investigated and the mechanisms involved were explored. Hyperlipidemia increased serum hepcidin, which was inhibited by TMP intervention. The results also indicated that TMP may decrease hepcidin expression via inhibition of Stat3 signaling. These findings suggest a promising rationale to prevent and hyperlidemia by targeting hepcidin or its upstream regulators, and highlight the potential application of natural compounds in treating hepcidin disorder­associated diseases.

The Influence of Hyperlipidemia on Endothelial Function of FPN1 Tek-Cre Mice and the Intervention Effect of Tetramethylpyrazine.

BACKGROUND/AIMS: Systemic iron homeostasis is strictly governed in mammals; however, disordered iron metabolism (such as excess iron burden) is recognized as a risk factor for various types of diseases including AS (Atherosclerosis). The hepcidin-ferroportin axis plays the key role in regulation of iron homeostasis and modulation of this signaling could be a potential therapeutic strategy in the treatment of these diseases. TMP (Tetramethylpyrazine) has been reported to have therapeutical effect on AS. Here, we aimed to investigate the effect of iron overload under hyperlipidemia condition on the endothelial injury, inflammation and oxidative stress by employing FPN1 Tek-cre mouse model with or without TMP intervention. METHODS: Subjects for this study were 80 FPN1 Tek-cre mice and 40 C57BL/6 mice and we randomly divided them into six groups: Group N: C57BL/6 mice with normal diet, Group M: C57BL/6 mice with high-fat diet, Group FN: FPN1 Tek-cre mice with normal diet, Group FNT: FPN1 Tek-cre mice with normal diet and TMP injection, Group FM: FPN1 Tek-cre mice with high-fat diet, Group FMT: FPN1 Tek-cre mice with high-fat diet and TMP injection. After seven days of treatment, blood samples were obtained to detect the levels of blood lipids, Hepcidin, NO, ET-1, ROS, MDA, SOD, IL-1, IL-6 and TNF-α respectively. The liver and aorta were used for testing the lipid deposition by using hematoxylin and eosin(HE). RESULTS: Hyperlipidemia could cause iron overload in the aorta and increased serum hepcidin level, particularly in FPN1 Tek-cre mice, and can be reversed by TMP intervention. Knockout of Fpn1 induced increase of serum hepcidin, exacerbated endothelial dysfunction, oxidative stress and inflammatory response, particularly under hyperlipidemia condition. TMP intervention attenuated these processes. CONCLUSIONS: Our study signifies the potential application of certain natural compounds to ameliorating iron disorders induced by hyperlipidemia and protecting on endothelial function through modulation of hepcidin-ferroportin signaling.

[Correlation Research on Blood Glucose Fluctuation in Type 2 Diabetes Mellitus Patients and Vas- cular Endothelial Injury/Platelet Activation/PKCß1 Expression].

Objective To observe the correlation between blood glucose fluctuation in type 2 dia- betes mellitus ( T2DM) patients and vascular endothelial injury/platelet activation/protein kinase Cß1 (PKCpß1). Methods Capillary blood was collected from finger tips of 38 T2DM patients at 7 time points, i.e., before 3 meals, 2 h after 3 meals, 21:00 pm before sleep. The mean amplitude of plasma glucose excursions (MAGE) was calculated. The peripheral blood platelet aggregation rate (PAG) induced by a- denosine diphosphate (ADP) and platelet membrane protein level of CD62p were determined by platelet fluorescent aggregometer and flow cytometry respectively. HbAlc was measured by ion-exchange high- performance liquid chromatography. Serum levels of E-selectin, von Willebrand factor ( vWF), and PKCß1 were detected by ELISA. Meanwhile, liver and renal functions, blood lipids were also measured. Their blood pressure was measured and body mass index (BMI) calculated. By taking HbA1c as a moni- tored index for assessing long-term glucose control, MAGE as an indicator for assessing glucose fluctua- tion, the correlations between serum markers for vascular endothelial injury (levels of E-selectin and vWF)/platelet activation indices (PAG and CD62p expression) and PKCß1 level/MAGE respectively were analyzed using Pearson correlation analysis and multivariant Logistic regression. The correlations be- tween PKCß1 level and MAGE/HbA1 c were also analyzed. Results In simple correlation analysis, there were no significant correlations between age/BMI/course of disease/medical history/serum levels of E-se- lectin/vWF/PKCß1/PAG/CD62p expression and MAGE (P >0. 05). There were significant correlations be- tween vascular endothelial injury markers ( E-selectin and vWF)/platelet activation indicators ( PAG, CD62p expression) and MAGE (r =0. 468, 0. 609, 0. 451 , 0. 674; P <0. 01). There were significant corre- lations between PKCß1 and glucose assessment indicators (MAGE and HbA1c)/vascular endothelial inju- ry markers ( E-selectin and vWF) , platelet activation indicators ( PAG and CD62p expression) (r = 0. 643, 0. 705, 0. 394, 0. 665, 0. 441 , 0. 577; P <0. 01). Conclusion PKCß1 , the key regulatory gene of coronary artery disease with blood stasis syndrome, was closely related with the degree of vascular en- dothelial injury and aggregation level of platelet activation in T2DM patients with blood glucose fluctuation.

Effects of Panax quinquefolium saponin on phosphatidylinositol 3-kinase/serine threonine kinase pathway of neonatal rat myocardial cells subjected to hypoxia.

OBJECTIVE: To explore the effects of Panax Quinquefolium Saponin (PQS) on phosphatidylinositol 3-kinase/serine threonine kinase (PI3K/Akt) pathway of neonatal rat myocardial cells subjected to hypoxia. METHODS: Neonatal rat myocardial cells were cultured in vitro. After the myocardial cell injury was induced by hypoxia, the cells were randomized into 5 groups: the normal group, the model group, the positive control group (Ciclosporin A, 2 µ mol/L), the low-dose PQS group (PQSL, 25mg/L), and the high-dose PQS group (PQSH, 50 mg/L). Morphology and behavior of myocardial cells were observed under an inverted microscope. Apoptosis rate and lactate dehydrogenase (LDH) leakage rate of myocardial cells were determined by colorimetry. Mitochondrial transmembrane potential was assessed using a fluorexon laser. Phospho-glycogen synthase kinase (GSK)-3ß and phospho-Akt as well as cytochrome C were determined by Western blot RESULTS: LDH leakage in the Ciclosporin A group, PQSH group and PQSL group reduced progressively compared with the model group (P<0.05). Akt and GSK-3ß was strongly phosphorylated after treatment with Ciclosporin A and PQS compared with the model group (P<0.05, P<0.01). Compared with the model group (16.41±1.74; 35.28±6.30), both the integrated optical density of mitochondrial permeability transition pore (MPTP) and the mitochondrial transmembrane potential significantly increased in the PQSH group (42.74±2.12; 71.36±6.54) and the PQSL group (39.58±1.49; 66.99±5.45; P<0.05, P<0.01). However, the protein of cytochrome C outside the mitochondrion decreased in the PQSH group (273.66±14.61) and the PQSL group (259.62±17.31) compared with the model group (502.41±17.76; P<0.05). CONCLUSION: Through activation of the PI3K/Akt pathway and inhibition of the MPTP, PQS might protect the heart against ischemia injury and apoptosis of myocardial cells.

[The estrogen-like protective effect of ginsenoside Rb3 on oxidative stress and dysfunction of endothelial cells induced by oxidized low-density lipoprotein].

Ginsenoside Rb3 (GRb3) is one of the main components in plasma of Panax quinquefolius Saponin of stem and leaf (PQS), which can be into human plasma. Previous studies have found PQS has estrogen-like vascular protective effects. In the present study, we investigated the estrogen-like protective effect of GRb3 on oxidative stress and dysfunction of endothelial cells induced by oxidized low-density lipoprotein. The activities of SOD, NOS and the contents of MDA in the cell lysate were examined by enzyme method or spectrophotometry. The NO and ET-1 concentrations in the cell culture supernatant were measured by ELISA method. The iNOS and eNOS mRNA expression were measured by real time RT-PCR, while the phosphorylation levels of Akt was measured by Western blotting. The results showed that GRb3 could enhance the activity of SOD, reduce the content of MDA, increase the level of NOS, NO, ET-1 and iNOS mRNA expression while decrease the eNOS mRNA expression and the phosphorylation level of Akt. These effects were blocked by estrogen receptor antagonist ICI182780. GRb3 can play a role in protecting vascular endothelial cells by estrogen receptors, the protective mechanism is similar to 17-ß estrodiol.

The Expression of CD14(+)CD16(+) Monocyte Subpopulation in Coronary Heart Disease Patients with Blood Stasis Syndrome.

Blood stasis syndrome (BSS), a comprehensive pathological state, is one of the traditional Chinese medicine syndromes of coronary heart disease (CHD). In our previous study, we investigated that Fc γ RIIIA (also called CD14(+)CD16(+) monocyte subpopulation) is one of the differentially expressed genes related to CHD patients and its possible role in the atherosclerotic formation and plaque rupture. However, whether or not the deregulation of CD14(+)CD16(+) monocyte subpopulation expression is implicated in the pathogenesis of CHD patients with BSS has not yet been elucidated. In this study, we found that there was no significant difference between CHD patients with BSS and non-BSS in CD14(+)CD16(+) monocyte subpopulation at gene level. Moreover, the protein level of CD14(+)CD16(+) monocyte subpopulation in CHD patients with BSS was increased significantly when compared to the CHD patients with non-BSS. Additionally, the level of inflammatory cytokines downstream of CD14(+)CD16(+) monocyte subpopulation such as TNF- α and IL-1 in sera was much higher in CHD patients with BSS than that in CHD patients with non-BSS. Taken together, these results indicated that CD14(+)CD16(+) monocyte subpopulation was implicated in the pathogenesis of CHD patients with BSS, which may be one of the bases of the essence of BSS investigation.

Influence of high blood glucose fluctuation on endothelial function of type 2 diabetes mellitus rats and effects of Panax Quinquefolius Saponin of stem and leaf.

OBJECTIVE: To observe the influence of high blood glucose fluctuation on the endothelial function of type 2 diabetes mellitus (T2DM) rats and the effects of Panax Quinquefolius Saponin (PQS) of stem and leaf. METHODS: The T2DM model was induced by intraperitoneal injection of a small dose of streptozotocin (STZ, 35 mg/kg) plus high fat and high caloric laboratory chow. Then, diabetic rats were divided into steady high blood glucose (SHG) group and fluctuant high blood glucose (FHG) group according to fasting blood glucose coefficient of variation (FBG-CV), and then, the FHG group rats were divided into 4 groups according to the level of FBG-CV and fasting blood glucose: PQS 30 mg/(kg·d) group, PQS 60 mg/(kg·d) group, metformin hydrochloride control (MHC) group, and FHG control group, 10 in each group. Meanwhile, 10 rats without any treatment were used as normal control (NOR) group. Eight weeks later, the aortic arteries histology, plasma hepatocyte growth factor (HGF), and serum nitric oxide (NO), endothelin-1 (ET-1), tumor necrosis factor α (TNF-α), and soluble intercellular adhesion molecule 1 (sICAM-1) were measured. RESULTS: In comparison with the NOR group, the level of plasma HGF and serum NO, ET-1 and TNF-α, and sICAM-1 in SHG and FHG control groups were all significantly increased (P<0.01); in comparison with the SHG group, plasma HGF and serum NO, ET-1, TNF-α, and sICAM-1 in FHG group were all significantly increased further (P<0.01 or P<0.05); meanwhile, in comparison with the FHG control group, the level of plasma HGF and serum NO, ET-1, TNF-α, and sICAM-1 in PQS and MHC groups were all decreased significantly (P<0.01). However, comparison of the aortic arteries histology among groups showed no significant differences either before or after treatment. CONCLUSION: Blood glucose fluctuation could facilitate the development of vascular endothelial dysfunction in T2DM rats, while PQS could improve the endothelial function of T2DM rats with high blood glucose fluctuation, which may be related to its effects of relieving vessel stress, decreasing vasoconstrictor ET-1 production, preventing compensated increase of NO, and reducing inflammatory reaction.

Ischemic postconditioning through percutaneous transluminal coronary angioplasty in pigs: roles of PI3K activation.

BACKGROUND: Ischemic postconditioning (IPOC) has been suggested to reduce ischemic reperfusion injury. It remains unclear whether the activation of phosphatidylinositol 3 kinase (PI3K)/Akt is a causal mechanism in the cardioprotection afforded by IPOC, which was examined in the model of percutaneous transluminal coronary angioplasty (PTCA) minipigs. METHODS AND RESULTS: Minipigs underwent 45-min occlusion of the left anterior descending artery and 24-h reperfusion by PTCA. Postconditioning was elicited by three cycles of 30-s reperfusion followed by 30-s ischemia at the onset of reperfusion. Infarct size was determined by triphenyl tetrazolium chloride staining after 24-h reperfusion, and mRNA and protein expression levels of PI3K were ascertained by reverse transcriptase-PCR and western-blot analysis in biopsies. Infarct size was significantly reduced and myocardial PI3K (Akt and GSK-3ß) phosphorylation was significantly increased with IPOC treatment compared with ischemic reperfusion. The administration of the PI3K inhibitor wortmannin (30 µg/kg) attenuated the protection of IPOC in the infarct size and decreased the expression of Akt and GSK-3ß phosphorylation compared with IPOC. IPOC had no impact on mRNA expression of AKT and GSK-3ß. CONCLUSION: Our findings show that IPOC is capable of protecting the myocardium against IR injury in the PTCA minipig model. The PI3K/Akt-signaling pathway is involved in the cardioprotective effect of IPOC.

Correlation between Fc γ R III a and aortic atherosclerotic plaque destabilization in ApoE knockout mice and intervention effects of effective components of chuanxiong rhizome and red peony root.

OBJECTIVE: To explore the correlation between Fc γ RIII A (CD16A) and aortic atherosclerotic plaque destabilization in apoE knockout (apoE KO) mice and the intervention effects of effective components of chuanxiong rhizome and red peony root. METHODS: Eight 8-week-old male C57BL/6J mice were selected as the control group. Forty 8-week-old male apoE KO mice were randomly divided into the model group, apoE KO + intraperitoneal injection immunoglobulin group (IVIG), apoE KO + simvastatin group (Sm), apoE KO + high dosage of xiongshao capsule (XSC) group (XSCH), and apoE KO + low dosage of XSC group (XSCL), 8 mice in each group. Mice in the control group were put on a normal diet, and others were fed with a high-fat diet. After 10-week different interventions, monocyte CD16 expression was detected by flow cytometry, aortic matrix metalloproteinase-9 (MMP-9) mRNA expression was detected using reverse transcription polymerase chain reaction, and serum tumor necrosis factor (TNF)-α level was detected using enzyme-linked immunosorbent assay. RESULTS: Compared with the control group, monocyte CD16 expression, aortic MMP-9 mRNA expression, and serum TNF-α level in the model group increased obviously (P<0.01). Injections of apoE KO mice with intraperitoneal immunoglobulin during a 5-day period significantly reduced the monocyte CD16 expression, aortic MMP-9 mRNA expression, and serum TNF-α level (P<0.01 or 0.05) over a 10-week period of high-fat diet. Indices above in the Sm group, XSCH group, and XSCL group decreased in a different degree. Of them, the aortic MMP-9 mRNA expression in XSCH group was lower than that in Sm group (P<0.05) and the monocyte CD16 expression and serum TNF-α level showed no significant difference between XSCH group and Sm group (P>0.05). Correlation analyses suggested positive correlation between monocyte CD16 expression and aortic MMP-9 mRNA expression or serum TNF-α level in IVIG group, XSCH group, and XSCL group. CONCLUSIONS: FcγR III A mediates systemic inflammation in the progression of coronary heart disease with blood stasis syndrome. XSC could stabilize atherosclerotic plaque by suppressing inflammation and its target was relative with FcγRIII A.