Histone demethylase KDM4A overexpression improved the efficiency of corrected human tripronuclear zygote development.

Human zygotes are difficult to obtain for research because of limited resources and ethical debates. Corrected human tripronuclear (ch3PN) zygotes obtained by removal of the extra pronucleus from abnormally fertilized tripronuclear (3PN) zygotes are considered an alternative resource for basic scientific research. In the present study, eight-cell and blastocyst formation efficiency were significantly lower in both 3PN and ch3PN embryos than in normal fertilized (2PN) embryos, while histone H3 lysine 9 trimethylation (H3K9me3) levels were much higher. It was speculated that the aberrant H3K9me3 level detected in ch3PN embryos may be related to low developmental competence. Microinjection of 1000 ng/µl lysine-specific demethylase 4A (KDM4A) mRNA effectively reduced the H3K9me3 level and significantly increased the developmental competence of ch3PN embryos. The quality of ch3PN zygotes improved as the grading criteria, cell number and pluripotent expression significantly increased in response to KDM4A mRNA injection. Developmental genes related to zygotic genome activation (ZGA) were also upregulated. These results indicate that KDM4A activates the transcription of the ZGA program by enhancing the expression of related genes, promoting epigenetic modifications and regulating the developmental potential of ch3PN embryos. The present study will facilitate future studies of ch3PN embryos and could provide additional options for infertile couples.

Bis-isatin derivatives: design, synthesis, and biological activity evaluation as potent dimeric DJ-1 inhibitors.

The PARK7 gene (encode DJ-1 protein) was first discovered as an oncogene and later found to be a causative gene for autosomal recessive early onset Parkinson's disease. DJ-1 has been proposed as a potential therapeutic anticancer target due to its pivotal role in tumorigenesis and cancer progression. Based on the homodimer structure of DJ-1, a series of bis-isatin derivatives with different length linkers were designed, synthesized, and evaluated as dimeric inhibitors targeting DJ-1 homodimer. Among them, DM10 with alkylene chain of C10 displayed the most potent inhibitory activity against DJ-1 deglycase. We further demonstrated that DM10 bound covalently to the homodimer of DJ-1. In human cancer cell lines H1299, MDA-MB-231, BEL7402, and 786-O, DM10 (2.5-20 µM) inhibited the cell growth in a concentration-dependent manner showing better anticancer effects compared with the positive control drug STK793590. In nude mice bearing H1299 cell xenograft, intratumor injection of DM10 (15 mg/kg) produced significantly potent tumor growth inhibition when compared with that caused by STK793590 (30 mg/kg). Moreover, we found that DM10 could significantly enhance N-(4-hydroxyphenyl)retinamide-based apoptosis and erastin-based ferroptosis in H1299 cells. In conclusion, DM10 is identified as a potent inhibitor targeting DJ-1 homodimer with the potential as sensitizing agent for other anticancer drugs, which might provide synergistical therapeutic option for cancer treatment.

The C terminus of DJ-1 determines its homodimerization, MGO detoxification activity and suppression of ferroptosis.

DJ-1 is a multifunctional protein associated with cancers and autosomal early-onset Parkinson disease. Besides the well-documented antioxidative stress activity, recent studies show that DJ-1 has deglycation enzymatic activity and anti-ferroptosis effect. It has been shown that DJ-1 forms the homodimerization, which dictates its antioxidative stress activity. In this study, we investigated the relationship between the dimeric structure of DJ-1 and its newly reported activities. In HEK293T cells with Flag-tagged and Myc-tagged DJ-1 overexpression, we performed deletion mutations and point mutations, narrowed down the most critical motif at the C terminus. We found that the deletion mutation of the last three amino acids at the C terminus of DJ-1 (DJ-1 ΔC3) disrupted its homodimerization with the hydrophobic L187 residue being of great importance for DJ-1 homodimerization. In addition, the ability in methylglyoxal (MGO) detoxification and deglycation was almost abolished in the mutation of DJ-1 ΔC3 and point mutant L187E compared with wild-type DJ-1 (DJ-1 WT). We also showed the suppression of erastin-triggered ferroptosis in DJ-1-/- mouse embryonic fibroblast cells was abolished by ΔC3 and L187E, but partially diminished by V51C. Thus, our results demonstrate that the C terminus of DJ-1 is crucial for its homodimerization, deglycation activity, and suppression of ferroptosis.

The cyclin-dependent kinase inhibitor, JNJ-7706621, improves in vitro developmental competence of porcine parthenogenetic activation and somatic cell nuclear transfer embryos.

In this study we examined the effects of JNJ-7706621, a cyclin-dependent kinase inhibitor, on the in vitro growth of pig embryos that had been produced either by parthenogenetic activation (PA) or somatic cell nuclear transfer (SCNT). A significantly higher percentage of PA embryos reached the blastocyst stage by Day 7 after exposure to 10µM JNJ-7706621 for 4h compared with embryos exposed to 5µgmL-1 cytochalasin B for 4h (P<0.05). Similarly, the rate of Tyr15 phosphorylation of the complex of cyclin and p34cdc2 (CDK1) was significantly elevated in the JNJ-7706621-treated embryos compared with embryos exposed to cytochalasin B or non-treated controls (P<0.05). In contrast, Thr161 phosphorylation of CDK1 was significantly lower in the JNJ-7706621-treated group compared with the cytochalasin B-treated as well as the non-treated group (P<0.05). Similarly, the level of M-phase-promoting factor (MPF) in embryos was significantly lower in the JNJ-7706621-treated group compared with the cytochalasin B-treated and non-treated groups (P<0.05). In addition, more SCNT embryos reached the blastocyst stage after treatment with JNJ-7706621 than following exposure to cytochalasin B (P<0.05). In conclusion, these results reveal that exposure to 10µM JNJ-7706621 for 4h improves early development of PA and SCNT porcine embryos by suppressing the activity of CDK1 and a concomitant reduction in the level of MPF.

Quisinostat treatment improves histone acetylation and developmental competence of porcine somatic cell nuclear transfer embryos.

Abnormal epigenetic modifications are considered a main contributing factor to low cloning efficiency. In the present study, we explored the effects of quisinostat, a novel histone deacetylase inhibitor, on blastocyst formation rate in porcine somatic-cell nuclear transfer (SCNT) embryos, on acetylation of histone H3 lysine 9 (AcH3K9), and on expression of POU5F1 protein and apoptosis-related genes BAX and BCL2. Our results showed that treatment with 10 nM quisinostat for 24 hr significantly improved the development of reconstructed embryos compared to the untreated group (19.0 ± 1.6% vs. 10.2 ± 0.9%; p < 0.05). Quisinostat-treated SCNT embryos also possessed significantly increased AcH3K9 at the pseudo-pronuclear stage (p < 0.05), as well as improved immunostaining intensity for POU5F1 at the blastocyst stage (p < 0.05). While no statistical difference in BAX expression was observed, BCL2 transcript abundance was significantly different in the quisinostat-treated compared to the untreated control group. Of the 457 quisinostat-treated cloned embryos transferred into three surrogates, six fetuses developed from the one sow that became pregnant. These findings suggested that quisinostat can regulate gene expression and epigenetic modification, facilitating nuclear reprogramming, and subsequently improving the developmental competence of pig SCNT embryos and blastocyst quality.

CDK inhibitor SU9516 induces tetraploid blastocyst formation from parthenogenetically activated porcine embryos.

OBJECTIVE: To examine the effect of SU9516, a cyclin-dependent kinase inhibitor, on the induction of tetraploid blastocyst formation in porcine embryos by parthenogenetic activation. RESULTS: Karyotype analysis of blastocysts showed that in the SU9516-treatment group 56% were tetraploid, whereas in the cytochalasin B (CB) group 67% were diploid. The level of maturation-promoting factor (MPF) in stimulated embryos treated with 10 µM SU9516 for 4 h was lower than in embryos treated with CB group (103 vs. 131 pg/ml). The mRNA expression levels of Nanog significantly increased in SU9516-treated embryos than CB group. CONCLUSION: SU9516 can induce tetraploid blastocyst formation at high efficiency. SU9516 can significantly influence the in vitro developmental competence of porcine parthenogenetically activated embryos by influencing the level of MPF and the gene related apoptosis and pluripotency.

RepSox improves viability and regulates gene expression in rhesus monkey-pig interspecies cloned embryos.

OBJECTIVE: To investigate the effect of the small molecule, RepSox, on the expression of developmentally important genes and the pre-implantation development of rhesus monkey-pig interspecies somatic cell nuclear transfer (iSCNT) embryos. RESULTS: Rhesus monkey cells expressing the monomeric red fluorescent protein 1 which have a normal (42) chromosome complement, were used as donor cells to generate iSCNT embryos. RepSox increased the expression levels of the pluripotency-related genes, Oct4 and Nanog (p < 0.05), but not of Sox2 compared with untreated embryos at the 2-4-cell stage. Expression of the anti-apoptotic gene, Bcl2, and the pro-apoptotic gene Bax was also affected at the 2-4-cell stage. RepSox treatment also increased the immunostaining intensity of Oct4 at the blastocyst stage (p < 0.05). Although the blastocyst developmental rate was higher in the group treated with 25 µM RepSox for 24 h than in the untreated control group (2.4 vs. 1.2%, p > 0.05), this was not significant. CONCLUSION: RepSox can improve the developmental potential of rhesus monkey-pig iSCNT embryos by regulating the expression of pluripotency-related genes.

The developmental competence of oocytes parthenogenetically activated by an electric pulse and anisomycin treatment.

OBJECTIVE: The aim of this study was to investigate the developmental competence of oocytes parthenogenetically activated by an electric pulse (EP) and treated with anisomycin and to determine whether this method is applicable to somatic cell nuclear transfer (SCNT). RESULTS: Embryos derived from porcine oocytes parthenogenetically activated by an EP and treatment with 0.01 µg/mL anisomycin had a significantly improved in vitro developmental capacity. Furthermore, 66.6% of blastocysts derived from these embryos had a diploid karyotype. The blastocyst formation rate of cloned embryos was similar between oocytes activated by an EP and treated with 2 mM 6-dimethylaminopurine for 4 h and those activated by an EP and treated with 0.01 µg/mL anisomycin for 4 h. The level of maturation-promoting factor was significantly decreased in oocytes activated by an EP and treated with anisomycin. Finally, the mRNA expression levels of apoptosis-related genes (Bax and Bcl-2) and pluripotency-related genes (Oct4, Nanog, and Sox2) were checked by RT-PCR. CONCLUSION: Our results demonstrate that porcine oocyte activation via an EP in combination with anisomycin treatment can lead to a high blastocyst formation rate in parthenogenetic activation and SCNT experiments.

Parthenogenetic activation and somatic cell nuclear transfer of porcine oocytes activated by an electric pulse and AZD5438 treatment.

We examined the in vitro developmental competence of parthenogenetic activation (PA) oocytes activated by an electric pulse (EP) and treated with various concentrations of AZD5438 for 4 h. Treatment with 10 µM AZD5438 for 4 h significantly improved the blastocyst formation rate of PA oocytes in comparison with 0, 20, or 50 µM AZD5438 treatment (46.4% vs. 34.5%, 32.3%, and 24.0%, respectively; P 0.05). Furthermore, 66.67% of blastocysts derived from these AZD5438-treated PA oocytes had a diploid karyotype. The blastocyst formation rate of PA and somatic cell nuclear transfer (SCNT) embryos was similar between oocytes activated by an EP and treated with 2 mM 6-dimethylaminopurine for 4 h and those activated by an EP and treated with 10 µM AZD5438 for 4 h (11.11% vs. 13.40%, P > 0.05). In addition, the level of maturation-promoting factor (MPF) was significantly decreased in oocytes activated by an EP and treated with 10 µM AZD5438 for 4 h. Finally, the mRNA expression levels of apoptosis-related genes (Bax and Bcl-2) and pluripotency-related genes (Oct4, Nanog, and Sox2) were checked by RT-PCR; however, there were no differences between the AZD5438-treated and non-treated control groups. Our results demonstrate that porcine oocyte activation via an EP in combination with AZD5438 treatment can lead to a high blastocyst formation rate in PA and SCNT experiments.

PCI-24781 can improve in vitro and in vivo developmental capacity of pig somatic cell nuclear transfer embryos.

OBJECTIVE: To examine the effect of PCI-24781 (abexinostat) on the blastocyst formation rate in pig somatic cell nuclear transferred (SCNT) embryos and acetylation levels of the histone H3 lysine 9 and histone H4 lysine 12. RESULTS: Treatment with 0.5 nM PCI-24781 for 6 h significantly improved the development of cloned embryos, in comparison to the control group (25.3 vs. 10.5 %, P < 0.05). Furthermore, PCI-24781 treatment led to elevated acetylation of H3K9 and H4K12. TUNEL assay and Hoechst 33342 staining revealed that the percentage of apoptotic cells in blastocysts was significantly lower in PCI-24781-treated SCNT embryos than in untreated embryos. Also, PCI-24781-treated embryos were transferred into three surrogate sows, one of whom became pregnant and two fetuses developed. CONCLUSION: PCI-24781 improves nuclear reprogramming and the developmental potential of pig SCNT embryos.

Generation of large pig and bovine blastocysts by culturing in human induced pluripotent stem cell medium.

We investigated the effect of human induced pluripotent stem cell (hiPS) medium on porcine somatic cell nuclear transfer and bovine in vitro fertilized early blastocysts, in comparison with North Carolina State University (NCSU)-37 medium and in vitro culture (IVC)-II medium. After 2 days of culture, the diameter of the portion of the blastocyst that was extruded from the zona pellucid dramatically differed between porcine blastocysts cultured in hiPS medium and those cultured in NCSU-37 medium (221.47 ± 38.94 µm versus 481.87 ± 40.61 µm, P < 0.01). Moreover, the diameter of the portion of the blastocyst significantly differed between bovine blastocysts cultured in hiPS medium and those cultured in IVC-II medium (150.30 ± 29.49 µm versus 195.58 ± 41.59 µm, P < 0.01). Furthermore, the total number of cells per porcine and bovine blastocyst was more than two-fold higher in blastocysts cultured in hiPS medium than in those cultured in NCSU-37 medium (44.33 ± 5.28 and 143.33 ± 16.05, P < 0.01) or IVC-II medium (172.12 ± 45.08 and 604.83 ± 242.64, P < 0.01), respectively. These results indicate that hiPS medium markedly improves the quality of porcine and bovine blastocysts.

PXD101 significantly improves nuclear reprogramming and the in vitro developmental competence of porcine SCNT embryos.

In this study, we investigated the effects of the histone deacetylase inhibitor PXD101 (belinostat) on the preimplantation development of porcine somatic cell nuclear transfer (SCNT) embryos and their expression of the epigenetic markers histone H3 acetylated at lysine 9 (AcH3K9). We compared the in vitro developmental competence of SCNT embryos treated with various concentrations of PXD101 for 24h. Treatment with 0.5 µM PXD101 significantly increased the proportion of SCNT embryos that reached the blastocyst stage, in comparison to the control group (23.3% vs. 11.5%, P<0.05). We tested the in vitro developmental competence of SCNT embryos treated with 0.5 µM PXD101 for various amounts of times following activation. Treatment for 24h significantly improved the development of porcine SCNT embryos, with a significantly higher proportion of embryos reaching the blastocyst stage in comparison to the control group (25.7% vs. 10.6%, P<0.05). PXD101-treated SCNT embryos were transferred into two surrogate sows, one of whom became pregnant and four fetuses developed. PXD101 treatment significantly increased the fluorescence intensity of immunostaining for AcH3K9 in embryos at the pseudo-pronuclear and 2-cell stages. At these stages, the fluorescence intensities of immunostaining for AcH3K9 were significantly higher in PXD101-treated embryos than in control untreated embryos. In conclusion, this study demonstrates that PXD101 can significantly improve the in vitro and in vivo developmental competence of porcine SCNT embryos and can enhance their nuclear reprogramming.

Production of Cloned Wuzhishan Miniature Pigs and Application for Alloxan Toxicity Test.

Wuzhishan miniature pig is one of the four most important pig breeds in China and has many major economic characteristics. Herein, we successfully used SCNT to clone Wuzhishan miniature pig. First, ear fibroblasts were isolated from a 2-year-old female Wuzhishan miniature piglet to be used as the donor cell. Second, good-quality COCs were selected from ovaries obtained from pigs at a local slaughterhouse and cultured. Mature eggs with the first polar body and ear fibroblasts were applied SCNT. Lastly, we in total produced 12 piglets with 7 piglets surviving to adults. Next, we used these pigs to test alloxan toxicity and to build T I D diabetes type. We know that diabetes mellitus is a chronic heterogeneous metabolic disease characterized by a high blood glucose level and abnormal insulin secretion. In this study, T I D (type I diabetes) was experimentally induced in cloned Wuzhishan miniature pigs with alloxan. In brief, an intravenous injection of alloxan (group B: 170 mg/kg, n = 3) was administered to pigs weighing between 27 and 39 kg. Sterile saline was administered to control pigs (n = 3). We determined the glycometabolism related index, performed an intravenous glucose tolerance test, and carried out immunohistochemistry experiments. There were no significant differences in body weight, blood glucose, and serum insulin in all groups, before treatment. The level of blood glucose was significantly higher (P < 0.05) in group B (12.18 ± 0.70 mmol/L) than in the control (2.93 ± 0.39 mmol/L). By contrast, the level of serum insulin was lower in group B (5.641 ± 0.573 µIU/mL) than in the control (7.578 ± 0.539 µIU/mL). Histological studies by hematoxylin and eosin (H&E) revealed a loss of ß-cells in the pancreas from pigs treated with 170 mg/kg alloxan. Immunolocalization studies showed a decrease in insulin reactivity in this treatment group as well. To conclude, our model holds promise in future studies of diabetes drug testing and islet xenotransplantation.

Production of rhesus monkey cloned embryos expressing monomeric red fluorescent protein by interspecies somatic cell nuclear transfer.

Interspecies somatic cell nuclear transfer (iSCNT) is a promising method to clone endangered animals from which oocytes are difficult to obtain. Monomeric red fluorescent protein 1 (mRFP1) is an excellent selection marker for transgenically modified cloned embryos during somatic cell nuclear transfer (SCNT). In this study, mRFP-expressing rhesus monkey cells or porcine cells were transferred into enucleated porcine oocytes to generate iSCNT and SCNT embryos, respectively. The development of these embryos was studied in vitro. The percentage of embryos that underwent cleavage did not significantly differ between iSCNT and SCNT embryos (P>0.05; 71.53% vs. 80.30%). However, significantly fewer iSCNT embryos than SCNT embryos reached the blastocyst stage (2.04% vs. 10.19%, P<0.05). Valproic acid was used in an attempt to increase the percentage of iSCNT embryos that developed to the blastocyst stage. However, the percentages of embryos that underwent cleavage and reached the blastocyst stage were similar between untreated iSCNT embryos and iSCNT embryos treated with 2mM valproic acid for 24h (72.12% vs. 70.83% and 2.67% vs. 2.35%, respectively). These data suggest that porcine-rhesus monkey interspecies embryos can be generated that efficiently express mRFP1. However, a significantly lower proportion of iSCNT embryos than SCNT embryos reach the blastocyst stage. Valproic acid does not increase the percentage of porcine-rhesus monkey iSCNT embryos that reach the blastocyst stage. The mechanisms underling nuclear reprogramming and epigenetic modifications in iSCNT need to be investigated further.

The relationship between serum FSH level and ovarian response during controlled ovarian stimulation.

OBJECTIVES: To evaluate whether serum follicle stimulating hormone (FSH) level during the early controlled ovarian stimulation can be used as a predictor of the ovarian response in the in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI) cycles. MATERIAL AND METHODS: The participants of this retrospective study were chosen from Reproductive Medicine Center, Weifang People's Hospital between January 2015 and December 2020.The participants of this study met the age of 20~43 years old, anti-Müllerian hormone (AMH) ≥ 1.2 ng/mL, antral follicle count (AFC) ≥ 5, and the data was complete and no cancellation cycle. Each participant was given GnRH agonist protocol and given a fixed dose of recombinant FSH in the first four days during the controlled ovarian stimulation (COS). According to the number of oocytes retrieved, the participants were divided into two different ovarian response groups. Serum FSH level after the fourth recombinant follicle stimulating hormone (rFSH) injection were compared during the different ovarian responders. RESULTS: The number of participants who met both the inclusion criteria and exclusion criteria was 235. Serum sFSH levels (mean: 11.76 ± 3.10 IU/L) in the inappropriate responders was significantly higher than serum sFSH levels (mean: 10.79 ± 2.52 IU/L) in the superior responders(p = 0.029). There was a weak correlation between serum sFSH levels and the number of oocytes retrieved (r = -0.134, p = 0.041). Serum sFSH levels had significant clinical valuable (p = 0.0346) in predicting the number of oocytes retrieved. CONCLUSIONS: Serum sFSH levels may be a potential marker to predict the ovarian response during the early COS in the IVF/ICSI cycles, which can guide the adjustment of the exogenous rFSH dose.

[Technological advances in single-cell genomic analyses].

The technological progress of the genomics has transformed life science research. The main objectives of genomics are sequencing of new genomes and genome-wide identification of the function and the interaction of genes and their products. The recently developed second generation or next generation sequencing platforms and DNA microarray technology are immensely important and powerful tools for functional genomic analyses. However, their application is limited by the requirement of sufficient amounts of high quality nucleic acid samples. Therefore, when only a single cell or a very small number of cells are available or are preferred, the whole genomic sequencing or functional genomic objectives cannot be achieved conventionally and require a robust amplification method. This review highlights DNA amplification technologies and summarizes the strategies currently utilized for whole genome sequencing of a single cell, with specific focus on studies investigating microorganisms; An outline for targeted re-sequencing enabling the analysis of larger genomes is also provided. Furthermore, the review presents the emerging functional genomic applications using next-generation sequencing or microarray analysis to examine genome-wide transcriptional profile, chromatin modification and other types of protein-DNA binding profile, and CpG methylation mapping in a single cell or a very low quantity of cells. The nature of these technologies and their prospects are also addressed.

Melatonin protects against oxybenzone-induced deterioration of mouse oocytes during maturation.

Oxybenzone (OBZ), an ultraviolet light filter that is widely used in sunscreens and cosmetics, is an emerging contaminant found in humans and the environment. Recent studies have shown that OBZ has been detected in women's plasma, urine, and breast milk. However, the effects of OBZ exposure on oocyte meiosis have not been addressed. In this study, we investigated the detrimental effects of OBZ on oocyte maturation and the protective roles of melatonin (MT) in OBZ-exposed mouse models. Our in vitro and in vivo results showed that OBZ suppressed oocyte maturation, while MT attenuated the meiotic defects induced by OBZ. In addition, OBZ facilitated H3K4 demethylation by increasing the expression of the Kdm5 family of genes, elevating ROS levels, decreasing GSH, impairing mitochondrial quality, and disrupting spindle configuration in oocytes. However, MT treatment resulted in significant protection against OBZ-induced damage during oocyte maturation and improved oocyte quality. The mechanisms underlying the beneficial roles of MT involved reduction of oxidative stress, inhibition of apoptosis, restoration of abnormal spindle assembly and up-regulation of H3K4me3. Collectively, our results suggest that MT protects against defects induced by OBZ during mouse oocyte maturation in vitro and in vivo.

[Yin-yang balance penetrating acupuncture combined with rehabilitation training on upper limb spasticity in stroke hemiplegia].

OBJECTIVE: To compare the therapeutic effect of yin-yang balance penetrating acupuncture combined with rehabilitation training and single rehabilitation training on upper limb spasticity in patients with stroke hemiplegia. METHODS: A total of 60 patients with upper limb spasticity of stroke hemiplegia were randomized into an observation group and a control group, 30 cases in each one. On the basis of conventional western medication, rehabilitation training was adopted in the control group. On the basis of treatment in the control group, yin-yang balance penetrating acupuncture was applied from Jianyu (LI 15) to Binao (LI 14), Quchi (LI 11) to Shaohai (HT 3), Yanglingquan (GB 34) to Yinlingquan (SP 9), Xuanzhong (GB 39) to Sanyinjiao (SP 6), etc. of the affected side in the observation group. The treatment was given once a day, 5 days were as one course, with a 2-day interval between two courses, 4 courses were required in both groups. The classification of modified Ashworth spasticity scale (MAS), surface integrated electromyogram (iEMG) of affected upper limb and the scores of National Institute of Health stroke scale (NIHSS), Fugl-Meyer assessment (FMA) of upper limb and modified Barthel index (MBI) before and after treatment were observed, the therapeutic effect was evaluated in both groups. RESULTS: ①After treatment, the MAS classification reduced in both groups (P<0.05), the cases of grade 0 to Ⅰ+ in the observation group were more than those in the control group (P<0.05); iEMG values of the maximum isometric voluntary contraction of affected usculus biceps brachii, musculus triceps brachii, musculus flexor carpi, musculus extensor carpi, extensor digitorum, aductor pollicis brevis were increased in both groups (P<0.05), and the variations of iEMG of above muscles on the affected side in the observation group were larger than those in the control group (P<0.05). ②After treatment, the scores of NIHSS were decreased (P<0.05), the scores of FMA, MBI were increased in both groups (P<0.05), and the variations of NIHSS, FMA and MBI scores were larger than those in the control group (P<0.05). ③The total effective rate was 93.3% (28/30) in the observation group, which was superior to 70.0% (21/30) in the control group (P<0.05). CONCLUSION: Yin-yang balance penetrating acupuncture combined with rehabilitation training can improve upper limb spasticity, heighten the motor function of upper limb and daily self care in patients with stroke hemiplegia, its therapeutic effect is superior to single rehabilitation training.

Comparative Proteomics Analysis Between the Short-Term Stress and Long-Term Adaptation of the Blattella germanica (Blattodea: Blattellidae) in Response to Beta-Cypermethrin.

A proteomic method combining two-dimensional polyacrylamide gel electrophoresis and tandem mass spectrometry was used to compare the hemolymph expression profiles of a beta-cypermethrin-resistant Blattella germanica L. strain (R) and a susceptible strain (S) after 24 h of beta-cypermethrin induction. The results showed that there were 42 differentially expressed proteins after induction of the R strain: 4 proteins were upregulated and 38 proteins were downregulated. One hundred one hemolymph proteins were differentially expressed after induction of the S strain: 53 proteins were upregulated and 48 proteins were downregulated. The identified proteins were mainly classified into the following categories: energy metabolism proteins such as arginine kinase and triose phosphate isomerase, detoxification-related proteins such as glutathione S-transferases (GSTs), signal molecule-regulated proteins such as nitric oxide synthase (NOS), and other proteins such as kinetic-related proteins and gene expression-related proteins. Several proteins show significant differences in response to short-term stress and long-term adaptation, and differential expression of these proteins reflects an overall change in cellular structure and metabolism associated with resistance to pyrethroid insecticides. In summary, our research has improved the understanding of the molecular mechanisms of beta-cypermethrin resistance in German cockroaches, which will facilitate the development of rational methods to improve the management of this pest.