Nutrition strategies to control post-weaning diarrhea of piglets: From the perspective of feeds.

Post-weaning diarrhea (PWD) is a globally significant threat to the swine industry. Historically, antibiotics as well as high doses of zinc oxide and copper sulfate have been commonly used to control PWD. However, the development of bacterial resistance and environmental pollution have created an interest in alternative strategies. In recent years, the research surrounding these alternative strategies and the mechanisms of piglet diarrhea has been continually updated. Mechanically, diarrhea in piglets is a result of an imbalance in intestinal fluid and electrolyte absorption and secretion. In general, enterotoxigenic Escherichia coli (ETEC) and diarrheal viruses are known to cause an imbalance in the absorption and secretion of intestinal fluids and electrolytes in piglets, resulting in diarrhea when Cl- secretion-driven fluid secretion surpasses absorptive capacity. From a perspective of feedstuffs, factors that contribute to imbalances in fluid absorption and secretion in the intestines of weaned piglets include high levels of crude protein (CP), stimulation by certain antigenic proteins, high acid-binding capacity (ABC), and contamination with deoxynivalenol (DON) in the diet. In response, efforts to reduce CP levels in diets, select feedstuffs with lower ABC values, and process feedstuffs using physical, chemical, and biological approaches are important strategies for alleviating PWD in piglets. Additionally, the diet supplementation with additives such as vitamins and natural products can also play a role in reducing the diarrhea incidence in weaned piglets. Here, we examine the mechanisms of absorption and secretion of intestinal fluids and electrolytes in piglets, summarize nutritional strategies to control PWD in piglets from the perspective of feeds, and provide new insights towards future research directions.

Molecular Insights to the Structure-Interaction Relationships of Human Proton-Coupled Oligopeptide Transporters (PepTs).

Human proton-coupled oligopeptide transporters (PepTs) are important membrane influx transporters that facilitate the cellular uptake of many drugs including ACE inhibitors and antibiotics. PepTs mediate the absorption of di- and tri-peptides from dietary proteins or gastrointestinal secretions, facilitate the reabsorption of peptide-bound amino acids in the kidney, and regulate neuropeptide homeostasis in extracellular fluids. PepT1 and PepT2 have been the most intensively investigated of all PepT isoforms. Modulating the interactions of PepTs and their drug substrates could influence treatment outcomes and adverse effects with certain therapies. In recent studies, topology models and protein structures of PepTs have been developed. The aim of this review was to summarise the current knowledge regarding structure-interaction relationships (SIRs) of PepTs and their substrates as well as the potential applications of this information in therapeutic optimisation and drug development. Such information may provide insights into the efficacy of PepT drug substrates in patients, mechanisms of drug-drug/food interactions and the potential role of PepTs targeting in drug design and development strategies.

Surgical Treatment of Severe Sprengel's Deformity: A Case Report.

CASE: An adolescent girl who presented with obviously impaired shoulder abduction due to untreated severe Sprengel's deformity underwent deformity correction surgery. Intraoperative neuromonitoring was used to warn of potential brachial plexus injury during a modified Woodward procedure. At 3-month follow-up, range of shoulder abduction had improved significantly. CONCLUSION: Sprengel's deformity is a rare congenital shoulder deformity, and the Woodward procedure could cause nerve injury in patients with severe Sprengel's deformity. Neuromonitoring can be performed intraoperatively to avoid brachial plexus injury.

Dietary supplementation with potassium-magnesium sulfate modulates the antioxidant capacity, immunity, and gut microbiota in weaned piglets.

The purpose of this study was to evaluate the effects of different levels of potassium magnesium sulfateon (PMS) on growth performance, diarrhea rate, intestinal morphology, antioxidant capacity, intestinal immunity, and gut microbiota in weaned piglets. A total of 216 weaned piglets were randomly divided into six dietary groups: the basal diet with 0% (CON), 0.15, 0.3, 0.45, 0.6, and 0.75% PMS. The results showed that the ADFI of 29-42 days and 1-42 days was linearly and quadratically increased by the PMS supplementation (P < 0.05), and significantly reduced the diarrhea rate in weaned piglets (P < 0.05). Moreover, dietary supplementation with PMS significantly reduced the serum adrenaline and noradrenaline levels in weaned piglets (P < 0.05). Furthermore, 0.3% PMS significantly increased the activity of glutathione peroxidase (GSH-Px) in the jejunum (P < 0.05) and tended to increase the activity of superoxide dismutase (SOD) in the jejunal mucosa of piglets (P < 0.1). Additionally, dietary supplementation with PMS significantly reduced the interleukin-1ß (IL-1ß) level in the jejunal mucosa (P < 0.05), and 0.3% PMS increased the serum IgM content in piglets (P < 0.05). Furthermore, the analysis of colonic microbiota by 16S RNA sequencing showed that the addition of PMS increased the Shannon index (P < 0.05) and Observed Species index (P < 0.05). Based on linear discriminant analysis effect size (LEfSe) and T-test analysis, the addition of PMS increased the relative abundance of Ruminococcaceae and Peptostreptococcaceae in the colonic digesta (P < 0.05). Spearman analysis showed that there was a positive correlation between intestinal GSH-Px activity and the relative abundance of Peptostreptococcaceae. These results showed that dietary supplementation with PMS could improve growth performance, alleviate diarrhea incidence, and modulate the antioxidant capacity and intestinal immunity in weaned piglets, which was partially related to the significant changes in colonic microbiota composition.

Lactobacillus reuteri 1 Enhances Intestinal Epithelial Barrier Function and Alleviates the Inflammatory Response Induced by Enterotoxigenic Escherichia coli K88 via Suppressing the MLCK Signaling Pathway in IPEC-J2 Cells.

Intestinal epithelial barrier injury disrupts immune homeostasis and leads to many intestinal disorders. Lactobacillus reuteri (L. reuteri) strains can influence immune system development and intestinal function. However, the underlying mechanisms of L. reuteri LR1 that regulate inflammatory response and intestinal integrity are still unknown. The present study aimed to determine the effects of LR1 on the ETEC K88-induced intestinal epithelial injury on the inflammatory response, intestinal epithelial barrier function, and the MLCK signal pathway and its underlying mechanism. Here, we showed that the 1 × 109 cfu/ml LR1 treatment for 4 h dramatically decreased interleukin-8 (IL-8) and IL-6 expression. Then, the data indicated that the 1 × 108 cfu/ml ETEC K88 treatment for 4 h dramatically enhanced IL-8, IL-6, and tumor necrosis factor-α (TNF-α) expression. Furthermore, scanning electron microscope (SEM) data indicated that pretreatment with LR1 inhibited the ETEC K88 that adhered on IPEC-J2 and alleviated the scratch injury of IPEC J2 cells. Moreover, LR1 pretreatment significantly reversed the declined transepithelial electrical resistance (TER) and tight junction protein level, and enhanced the induction by ETEC K88 treatment. Additionally, LR1 pretreatment dramatically declined IL-8, IL-17A, IL-6, and TNF-α levels compared with the ETEC K88 group. Then, ETEC K88-treated IPEC-J2 cells had a higher level of myosin light-chain kinase (MLCK), higher MLC levels, and a lower Rho-associated kinase (ROCK) level than the control group, while LR1 pretreatment significantly declined the MLCK and MLC expression and enhanced ROCK level in the ETEC K88-challenged IPEC-J2 cells. Mechanistically, depletion of MLCK significantly declined MLC expression in IPEC-J2 challenged with ETEC K88 compared to the si NC+ETEC K88 group. On the other hand, the TER of the si MLCK+ETEC K88 group was higher and the FD4 flux in the si MLCK+ETEC K88 group was lower compared with the si NC+ETEC K88 group. In addition, depletion of MLCK significantly enhanced Claudin-1 level and declined IL-8 and TNF-α levels in IPEC-J2 pretreated with LR1 followed by challenging with ETEC K88. In conclusion, our work indicated that L. reuteri LR1 can decline inflammatory response and improve intestinal epithelial barrier function through suppressing the MLCK signal pathway in the ETEC K88-challenged IPEC-J2.

Utilization of intraoperative neuromonitoring during the Woodward procedure for treatment of Sprengel deformity.

BACKGROUND: Brachial plexus injury (BPI) leading to palsy of the upper extremities is the most serious complication of the Woodward procedure for treatment of Sprengel deformity. Intraoperative neuromonitoring (IONM) is widely used for detecting emerging spinal cord or peripheral nerve injury during spinal and shoulder surgery. However, to date, its utilization in pediatric patients with Sprengel deformity is limited. Furthermore, it remains unclear whether IONM can help prevent BPI during surgery. The purpose of the current study was to assess the feasibility and effectiveness of IONM for early identification and prevention of nerve injury during the Woodward procedure. METHODS: We retrospectively reviewed the records of patients who underwent the Woodward procedure for Sprengel deformity at our institution between January 2017 and January 2020. IONM, including somatosensory evoked potentials (SEP) and motor evoked potentials (MEPs), was performed in all patients. Detailed IONM data were collected and analyzed. Preoperative and postoperative cosmetic appearance (according to the Cavendish classification), shoulder joint abduction function, and radiologic evaluation of the scapula were reviewed. Surgical complications were recorded. RESULTS: Forty-six patients (19 girls, 27 boys) were included (mean age, 5.1 ± 2.1 years). Both SEP and MEP (amplitude of the abductor pollicis) were successfully performed (100%). MEP alerts occurred in 3 patients (6.5%). After scapula position adjustment, signals recovered in 2 patients and remained unchanged in 1 patient-this patient exhibited postoperative motor deficits that resolved completely by 4 months recovery. The SEP amplitudes decreased in all 3 patients but did not reach the warning criteria. Forty patients were classified as grade III and 6 as grade IV in the Cavendish classification, whereas 35 patients were classified as grade II and 11 as grade III in the Rigault scale. The preoperative Cavendish grade was III (III, IV) and the postoperative Cavendish grade was I (I, II) (χ2 = 88.098, P < .001). The preoperative Rigault grade was II (II, III) and the postoperative Rigault grade was I (I, II) (χ2 = 62.133, P < .001). The mean arc of shoulder joint abduction improved from 99° ± 8° to 167° ± 7° (t = -45.871, P < .001) after surgery. Except for temporary motor deficits detected in 1 patient, no other postoperative complications were observed through the time of final follow-up. CONCLUSION: IONM during the Woodward procedure for Sprengel deformity is feasible and effective in detecting intraoperative neurologic changes and may be effective in preventing BPI associated with surgery.

Intestinal morphology and immune profiles are altered in piglets by early-weaning.

This study was to investigate the developmental changes in intestinal morphology and immune profiles in suckling and weaning piglets. Seventy-two weaning piglets with equal initial body weight from 8 litters (Duroc × Landrace × Yorkshire, 9 piglets per litter) were selected. Thirty-two piglets in the suckling group were nursed by sows until they were 17, 21, 28, or 35 days of age. While the other forty piglets were weaned at 14 d of age, and then housed in the same farrowing cage without a sow and slaughtered until they were on d 0, 3, 7, 14 and 21 after weaning at d 14 of age (wd 0, 3, 7, 14, 21). Blood, jejunal mucosa, intraepithelial lymphocyte (IEL) and lamina propria T lymphocyte (LPL) were harvested from suckling piglets at d 14, 17, 21, 28 and 35 of age and weaning piglets on d 0, 3, 7, 14 and wd 0, wd 3, wd 7, wd 14 and wd 21). The results showed that compared with the wd 0, early weaning significantly declined the average daily gain of postweaning 0-7 (wd 0-7) (P < 0.05), and jejunal villus height on wd 3 (P < 0.05), as well as increased the jejunal crypt depth of piglets on wd 7, 14 and 21 (P < 0.05). And there were no significant differences in average daily gain and villus height after suckling (P > 0.05). The level of serum interferon-γ (IFN-γ) was increased on wd 7 and decreased on wd 21 (P < 0.05), while IFN-γ level in jejunal mucosa was enhanced at wd 3 in comparison with wd 0 (P < 0.05). And the serum interleukin-4 (IL-4) levels were increased at wd 14, wd 21, but the mucosa IL-4 concentration was strikingly increased on wd 7 (P < 0.05). Moreover, weaning led to the enhanced levels of interleukin-1b (IL-1b), interleukin-2 (IL-2) and soluble interleukin-2 receptor (sIL-2R) in serum (P < 0.05), as well as declined the levels of sIL-2R in jejunal mucosa at wd 3 (P < 0.05). In suckling piglets, the serum immunoglobulin A (IgA), IL-4 and IL-2 levels were increased with increasing age (P < 0.05), whereas the jejunal mucosa IL-1b and serum sIL-2R levels were lower (P < 0.05). Furthermore, significantly lower CD4 percentage in peripheral blood T lymphocyte subsets were found at wd 3 and wd 7 (P < 0.05), whereas the CD8 percentage in peripheral blood T lymphocyte subsets were enhanced on wd 3 and wd 7 than wd 0 (P < 0.05). Moreover, the weaning piglets at wd 3 had a lower CD4/CD8 ratio than wd 0 (P < 0.05). Additionally, we found that weaning decreased IgG, IL-4, IL-2 and IL-1b levels of IEL during 1-week post-weaning (P < 0.05). Similarly, the levels of IgA, IgG, IL-2 and sIL-2R in LPL medium were also declined from piglets postweaning 1 week (P < 0.05). Early weaning reduced the growth performance, damaged jejunal morphology, disrupted IFN-γ/IL-4, IL-2/sIL-2R and T lymphocyte balance, and impaired the IEL and LPL immune profiles of piglets.

AMPK-PINK1/Parkin Mediated Mitophagy Is Necessary for Alleviating Oxidative Stress-Induced Intestinal Epithelial Barrier Damage and Mitochondrial Energy Metabolism Dysfunction in IPEC-J2.

The imbalance of redox biology and oxidative stress leads to intestinal barrier injury and mitophagy. However, much uncertainty still exists about the role of mitophagy in oxidative stress and intestinal function. Here, we showed the effects of hydrogen peroxide (H2O2)-induced oxidative stress on intestinal epithelial cell oxidation balance, intestinal barrier function and mitochondrial energy metabolism and its underlying mechanism. In this study, we found that H2O2-induced oxidative stress activated adenosine monophosphate-activated protein kinase (AMPK) and enhanced mitophagy in intestinal porcine epithelial cells (IPEC-J2). While compound C (AMPK inhibitor) and mdivi-1 (mitophagy inhibitor) significantly reduced the activity of superoxide dismutase (SOD) and increased mitochondrial reactive oxygen species (ROS) levels in H2O2 treated cells. Moreover, compound C and mdivi-1 significantly reduced the trans-epithelium electrical resistant (TER) and increased the fluorescein isothiocyanate-dextran (FD4) flux in H2O2 treated IPEC-J2. Furthermore, compound C and mdivi-1 significantly reduced the activity of mitochondrial complex II. Seahorse XF96 data showed that compound C + mdivi-1+ H2O2 treatment significantly reduced maximum respiratory oxygen consumption and spare respiratory capacity. Additionally, compound C or mdivi-1 treatment reduced the formation of mitochondrial autophagosomes. These results unveiled that AMPK and PINK1/Parkin mediated mitophagy is necessary for alleviating oxidative stress induced intestinal epithelial barrier damage and mitochondrial energy metabolism dysfunction in IPEC-J2.

Transcriptomic Analyses Reveal B-Cell Translocation Gene 2 as a Potential Therapeutic Target in Ovarian Cancer.

Ovarian cancer is the most common and aggressive type of tumor of the female reproductive system. Two factors account for this detrimental clinical presentation: (i) the lack of early detection methods and (ii) the inherently aggressive nature of this malignancy. Currently, transcriptomic analyses have become important tools to identify new targets in different cancer types. In this study, by measuring expression levels in ovarian cancer samples and stem cell samples, we identified 24 tumor suppressor genes consistently associated with poor prognosis. Combined results further revealed a potential therapeutic candidate, BTG2, which belongs to the antiproliferative gene family. Our results showed that BTG2 expression regulated ovarian cancer cell proliferation via G1/S phase cell cycle arrest by regulating Cyclin D1, CDK4, p-AKT, and p-ERK expression. BTG2 also inhibited cell migration by modulating MMP-2 and MMP-9 expression. Furthermore, xenograft models confirmed a growth inhibitory effect of BTG2 in ovarian cancer in vivo. BTG2 was significantly associated with ovarian cancer FIGO stage and grade in the clinic. Our findings indicated that BTG2 exerts a suppressive impact on ovarian cancer and could be a potential biomarker.

Scalp acupuncture enhances local brain regions functional activities and functional connections between cerebral hemispheres in acute ischemic stroke patients.

This study aimed to explore the changes in functional connections between cerebral hemispheres and local brain regions functional activities in patients with acute ischemic stroke (AIS) treated with International Standard Scalp Acupuncture (ISSA). Thirty patients with middle cerebral artery AIS in the dominant hemisphere were selected and randomly divided into two groups such as the control group and the scalp acupuncture group, with 15 patients in each group. Patients in the control group were treated with conventional Western medicine, while patients in the scalp acupuncture group received ISSA (acupuncture at the parietal midline [MS5], acupuncture at the left anterior parietotemporal oblique line [MS6] and acupuncture at the left posterior parietotemporal oblique line [MS7]) for one course of treatment. All patients were evaluated for treatment efficacy and received whole brain resting state functional magnetic resonance imaging (Rs-fMRI) scan before and after treatment. The observational indicators included: (a) the National Institutes of Health Stroke Scale (NIHSS) scores and the simplified Fugl-Meyer Assessment (SFMA) scores; (b) analyses of the amplitude of low-frequency fluctuation (ALFF), regional homogeneity (ReHo) and voxel-mirrored homotopic connectivity (VMHC). The results showed a significant difference in the NIHSS scores before and after treatment in the scalp acupuncture group compared with the control group (p < .05), indicating that patients improved better after scalp acupuncture treatment. Compared with the control group, the VMHC, ALFF and ReHo values in the scalp acupuncture group increased after treatment. The VMHC values increased in the brain regions dominated by bilateral BA6 and BA8; the ALFF values increased in the left BA39 and the adjacent superior temporal gyrus and middle temporal gyrus; and the ReHo values increased in the brain regions extending from left middle temporal gyrus (including BA21) to BA37, and the brain regions extending from the left BA40 and angular gyrus to BA7. The present study indicated that scalp acupuncture can specifically strengthen the functional activities of the brain regions related to sensory integration, language processing and motor coordination in the middle aged and elderly patients with AIS of the dominant cerebral hemisphere, and can strengthen bilateral frontal lobe motor control. This study may provide a scientific basis for the clinical application of ISSA treatment in patients with AIS, and may also provide a preliminary research basis for further animal experiments.

Hemispheric Difference of Regional Brain Function Exists in Patients With Acute Stroke in Different Cerebral Hemispheres: A Resting-State fMRI Study.

OBJECTIVE: To explore the different compensatory mechanisms of brain function between the patients with brain dysfunction after acute ischemic stroke (AIS) in the dominant hemisphere and the non-dominant hemisphere based on Resting-state Functional Magnetic Resonance Imaging (Rs-fMRI). METHODS: In this trial, 15 healthy subjects (HS) were used as blank controls. In total, 30 hemiplegic patients with middle cerebral artery acute infarction of different dominant hemispheres were divided into the dominant hemisphere group (DH) and the non-dominant hemisphere group (NDH), scanned by a 3.0 T MRI scanner, to obtain the amplitude of low-frequency fluctuations (ALFF) and regional homogeneity (ReHo) and compare the differences. RESULTS: Compared with the HS, increased ALFF values in the brain areas, such as the bilateral midbrain, were observed in DH. Meanwhile decreased ReHo values in the brain areas, such as the right postcentral gyrus (BA3), were also observed. Enhanced ALFF values in the brain areas, such as the left BA6, and enhanced ReHo values in the brain areas, such as the left precuneus, were observed in the NDH. The ALFF and ReHo values of the right BA9 and precentral gyrus were both increased. Compared with DH, the NDH group showed lower ALFF values in the left supplementary motor area and lower ReHo values in the right BA10. CONCLUSION: After acute infarction in the middle cerebral artery of the dominant hemisphere, a compensation mechanism is triggered in brain areas of the ipsilateral cortex regulating motor-related pathways, while some brain areas related to cognition, sensation, and motor in the contralateral cortex are suppressed, and the connection with the peripheral brain regions is weakened. After acute infarction in the middle cerebral artery of the non-dominant hemisphere, compensatory activation appears in motor control-related brain areas of the dominant hemisphere. After acute middle cerebral artery infarction in the dominant hemisphere, compared with the non-dominant hemisphere, functional specificity in the bilateral supplementary motor area weakens. After acute middle cerebral artery infarction in different hemispheres, there are hemispheric differences in the compensatory mechanism of brain function.

Phenethyl Isothiocyanate Enhances the Cytotoxic Effects of PARP Inhibitors in High-Grade Serous Ovarian Cancer Cells.

While PARP inhibitor (PARPi) therapies have shown promising results in the treatment of high-grade serous ovarian cancer (HGSOC) harboring homologous recombination deficiencies, primary resistance to PARPi frequently occurs and even initial responders may eventually become resistant. Therefore, the development of novel effective combinatorial strategies to treat HGSOC is urgently needed. Here, we report that H2O2-induced oxidative stress sensitized HGSOC cells to PARPi BMN 673. Furthermore, Phenethyl isothiocyanate (PEITC) as a ROS-inducing agent significantly enhanced the cytotoxic effects of BMN 673. Mechanistically, combined use of PEITC and BMN 673 resulted in ROS overproduction and accumulation, enhanced DNA damage, G2/M arrest and apoptosis, all of which were significantly reversed by the ROS scavenger N-Acetyl-L-cysteine. We also showed that while PEITC did not further enhance the ability of BMN 673 on PARP1 trapping in HGSOC cells, the therapeutic effects of the PEITC/BMN 673 combination were at least in part dependent on the presence of PARP1. Importantly, the PEITC/BMN 673 combination potently abrogated the growth of HGSOC tumor spheroids and patient-derived organoid models of HGSOC and cervical cancer. Our findings provide a basis for further investigation of the utility of PARPi combination regimen in HGSOC and cervical cancer through ROS-mediated mechanisms.

Modulatory effect of International Standard Scalp Acupuncture on brain activation in the elderly as revealed by resting-state fMRI.

The specific mechanisms by which acupuncture affects the central nervous system are unclear. In the International Standard Scalp Acupuncture system, acupuncture needles are applied at the middle line of the vertex, anterior parietal-temporal oblique line, and the posterior parietal-temporal oblique line. We conducted a single-arm prospective clinical trial in which seven healthy elderly volunteers (three men and four women; 50-70 years old) received International Standard Scalp Acupuncture at MS5 (the mid-sagittal line between Baihui (DU20) and Qianding (DU21)), the left MS6 (line joining Sishencong (EX-HN1) and Xuanli (GB6)), and the left MS7 (line joining DU20 and Qubin (GB7)). After acupuncture, resting-state functional magnetic resonance imaging demonstrated changes in the fractional amplitude of low frequency fluctuations and regional homogeneity in various areas, showing remarkable enhancement of regional homogeneity in the bilateral anterior cingulate, left medial frontal gyrus, supramarginal gyrus, right middle frontal gyrus, and inferior frontal gyrus. Functional connectivity based on a seed region at the right middle frontal gyrus (42, 51, 9) decreased at the bilateral medial superior frontal gyrus. Our data preliminarily indicates that the international standard scalp acupuncture in healthy elderly participants specifcally enhances the correlation between the brain regions involved in cognition and implementation of the brain network regulation system and the surrounding adjacent brain regions. The study was approved by the Ethics Committee of the China-Japan Union Hospital at Jilin University, China, on July 18, 2016 (approval No. 2016ks043).

Bafilomycin A1 induces caspase-independent cell death in hepatocellular carcinoma cells via targeting of autophagy and MAPK pathways.

Hepatocellular carcinoma (HCC) is refractory to chemotherapies, necessitating novel effective agents. The lysosome inhibitor Bafilomycin A1 (BafA1) at high concentrations displays cytotoxicity in a variety of cancers. Here we show that BafA1 at nanomolar concentrations suppresses HCC cell growth in both 2 dimensional (2D) and 3D cultures. BafA1 induced cell cycle arrest in the G1 phase and triggered Cyclin D1 turnover in HCC cells in a dual-specificity tyrosine phosphorylation-regulated kinase 1B (DYRK1B) dependent manner. Notably, BafA1 induced caspase-independent cell death in HCC cells by impairing autophagy flux as demonstrated by elevated LC3 conversion and p62/SQSTM1 levels. Moreover, genetic ablation of LC3 significantly attenuated BafA1-induced cytotoxicity of HCC cells. We further demonstrate that pharmacological down-regulation or genetic depletion of p38 MAPK decreased BafA1-induced cell death via abolishment of BafA1-induced upregulation of Puma. Notably, knockdown of Puma impaired BafA1-induced HCC cell death, and overexpression of Puma enhanced BafA1-mediated HCC cell death, suggesting a role for Puma in BafA1-mediated cytotoxicity. Interestingly, pharmacological inhibition of JNK with SP600125 enhanced BafA1-mediated cytotoxicity both in vitro and in xenografts derived from HCC cells. Taken together, our data suggest that BafA1 may offer potential as an effective therapy for HCC.

Effects of Temperature on the Termination of Egg Diapause and Post-Diapause Embryonic Development of Galeruca daurica (Coleoptera: Chrysomelidae).

Galeruca daurica (Joannis) is a new pest on the grasslands of Inner Mongolia, China. It is univoltine and overwinters in the egg stage. Larvae and adults feed on the foliage of Allium plants. To assess the requirements to terminate egg diapause and subsequent effects on post-diapause development rate, eggs were held at different temperature regimes. Exposure to low temperatures was required to terminate egg diapause. Prolonged exposure (2 mo vs 1 mo) to 5°C and outside ambient conditions (mean temperature: 10.5°C; range: -7.1-21.6°C) enhanced the termination of egg diapause. Prolonged exposure also reduced the time to egg hatch; e.g., eggs held for 2 mo versus 1 mo at 5°C developed more quickly when subsequently placed at warmer temperatures. Egg hatch was observed at 17, 21, 25, and 29°C, but not at 15°C. Regression analysis identified 16.2°C as the minimum temperature for post-diapause development. The temperature requirement to complete embryonic development (from diapause termination to egg hatch) was calculated to be 103.1 to 140.9 degree-days.

[Expression and clinical significance of Dyrk1b in the specimens and cells of cervical lesions].

OBJECTIVE: To detect and explore the expression and clinical significance of dual specificity tyrosine phosphorylation regulated kinase1b (Dyrk1b) in the specimens and cells of cervical lesions. METHODS: (1) All the data were collected from 75 patients with cervical cancer and 52 cases with squamous intraepithelial lesion (SIL) admitted in the First Affiliated Hospital of Dalian Medical College during Jan. 2011 to Dec. 2013 and confirmed by pathological examination, included 60 cases of stage Ⅰ and 15 cases of stage Ⅱ, 12 cases with low-grade squamous intraepithelial lesion (LSIL) and 40 cases with high-grade squamous intraepithelial lesion (HSIL). While, 28 cases with chronic cervicitis were chosen as the control group. The protein expression of Dyrk1b was detected by immunohistochemistry among the four groups. (2) The expression of Dyrk1b in HeLa and SiHa cells were detected by western blot method and the expression of Dyrk1b protein were also detected after treatment of AZ191 (5, 10 µmol/L) for 48 hours in HeLa and SiHa cells. (3) The cellular survival and proliferation of HeLa and SiHa cells treated by different concentrations of AZ191 (2.5, 5, 10, 25, 50, 100 µmol/L) for 48 hours were detected by methyl thiazolyl tetrazolium (MTT) assay. (4) The rate of apoptosis of HeLa and SiHa cells was detected by flowcytometry after treatment of AZ191 (5, 10 µmol/L) for 48 hours. RESULTS: (1) The positive rates of Dyrk1b protein in chronic cervicitis, LSIL, HSIL and cervical squamous cancer by immunohistochemistry were 11%(3/28), 1/12, 42% (17/40) and 71% (53/75), respectively. The expression of Dyrk1b in cervical squamous cancer and HISL were higher than those in LSIL and chronic cervicitis (P<0.01), there were significant difference between cervical squamous cancer and HSIL, or between HSIL and LSIL (all P<0.05), while there were not significant difference between LSIL and chronic cervicitis (P>0.05). Expression of Dyrk1b was correlated with stromal invasion depth of cervical cancer (P<0.05), but not with age, clinical stage, lymph node metastasis, and serum squamous cell carcinom antigen (SCC-Ag) levels (all P>0.05). (2) Dyrk1b protein was expressed in different levels in HeLa and SiHa cells, and the expression of Dyrk1b was decreased gradually as the increased of the concentration of AZ191 in both HeLa and SiHa cells by treatment of AZ191 for 48 hours. (3) Different concentration of AZ191 treated on cervical cancer cells could inhibit the cellular proliferation and induce cell apoptosis in a concentration-dependent manner (P<0.01), concomitant to the decreased cell survival rate. The apoptosis rate of HeLa and SiHa were increased significantly after 10 µmol/L AZ191-treatment for 48 hours, but no any difference induced by 5 µmol/L AZ191-treatment compared to control group. Also,there was no any difference between Hela and SiHa cells in either inhibitory effect or apoptosis rate induced by AZ191. CONCLUSIONS: Dyrk1b is over-expressed in either specimens or cells of cervical cancer. The expression of Dyrk1b protein in cervical lesions is increased as the progression of disease. Dyrk1b inhibitor AZ191 could inhibit cellular proliferation and induce apoptosis in a concentration-dependent manner in cervical cancer cells.

Mirk/Dyrk1B mediates G0/G1 to S phase cell cycle progression and cell survival involving MAPK/ERK signaling in human cancer cells.

BACKGROUND: Mirk/Dyrk1B contributes to G0 arrest by destabilization of cyclin D1 and stabilization of p27kip1 to maintain the viability of quiescent human cancer cells, and it could be negatively regulated by mitogenic-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) signaling. This study was performed to investigate the effect of Mirk/Dyrk1B on cell cycle and survival of human cancer cells involving MAPK/ERK signaling. METHODS: The correlations between Mirk/Dyrk1B expression and active ERK1/2 detected by western blot in both ovarian cancer and non-small cell lung cancer (NSCLC) cells were analyzed by simple regression. Mirk/Dyrk1B unique phosphopeptides with sites associated with Mirk/Dyrk1B protein were isolated and quantitated by liquid chromatography coupled to tandem mass/mass spectrometry (LC-MS/MS) proteomics analysis. The human cancer cells were treated with small interfering RNAs (siRNAs) and/or U0126, an inhibitor of MEK for indicated duration, followed by investigating the alterations of cell cycle and apoptosis as well as related proteins examined by flow cytometry and Western blot, respectively. RESULTS: Our study demonstrated the widely expressed Mirk/Dyrk1B proteins in the human cancer cells were positively correlated with the levels of activated ERK1/2. Moreover, Mirk/Dyrk1B protein expressions consistent with the tyrosine autophosphorylated levels in the human cancer cells were increased by U0126 or growth factor-depleted culture. Conversely, knockdown of Mirk/Dyrk1B by siRNA led to up-regulated activation of c-Raf-MEK-ERK1/2 pathway and subsequent changes in cell cycle proteins (cyclin D1, p27kip1), accompanied by increased growth rate and cells from G0/G1 into S of cell cycle which could be blocked by U0126 in a dose-dependent manner, indicating Mirk/Dyrk1B may sequester MAPK/ERK pathway, and vice versa. Whereas, combined Mirk siRNA and U0126 induced cell apoptosis in the human cancer cells. CONCLUSIONS: These data together show that Mirk/Dyrk1B mediates cell cycle and survival via interacting with MAPK/ERK signals and simultaneous inhibition of both pathways may be a novel therapeutic target for human cancer.

[Percutaneous vertebroplasty to treat osteoporotic vertebral compression fractures combined with intravertebral clefts by unilateral approach].

OBJECTIVE: To evaluate the effectiveness of percutaneous vertebroplasty (PVP) in the treatment of osteoporotic vertebral compression fractures with or without intravertebral clefts by unilateral approach and the impact of intravertebral clefts on the effectiveness. METHODS: The clinical data of 65 patients who met the inclusion criteria of osteoporotic vertebral compression fracture were retrospectively analyzed. According to having intravertebral clefts or not, the patients were divided into 2 groups: cleft group (group A, n=25) and non-cleft group (group B, n=40). There was no significant difference in gender, age, cause of injury, the level of fracture vertebrae, degree of damage, and interval of injury and operation between 2 groups (P > 0.05). All patients were given PVP procedure by unilateral approach. The operation time, the injected volume of bone cement, time to ambulate, complications, and adjacent vertebral re-fracture were recorded. The height of anterior and middle column and the posterior convex Cobb angle of injured spine were measured on the lateral X-ray film in standing position at preoperation and 1, 48 weeks after operation. The visual analogue scale (VAS) score and Oswestry disability index (ODI) system were used to evaluate the pain relief and improvement of daily activity function respectively at preoperation and 1, 4, and 48 weeks after operation. RESULTS: There was no significant difference in the operation time and time to ambulate between 2 groups (P > 0.05). The injected volume of bone cement in group B was significantly less than that in group A (t=1.833, P=0.034). Asymptomatic cement leakage occurred in 6 patients (4 in group A and 2 in group B), in group A including 1 case of venous leakage, 2 cases of paravertebral leakage, and 1 case of intradiscal leakage; in group B including 2 cases of venous leakage. No symptomatic pulmonary embolism was observed. The vital sign was stable during operation and postoperatively. All patients were followed up 12-30 months (mean, 18.5 months). No re-fracture of the vertebrae occurred during the follow-up. The postoperative VAS score, ODI, the height of anterior and middle column, and the posterior convex Cobb angle of injured spine were improved significantly when compared with the preoperative ones in 2 groups (P < 0.05), but no significant difference was found between 2 groups at pre- and post-operation (P > 0.05). CONCLUSION: PVP by unilateral approach is safty and efficacy in the treatment of osteoporosis vertebral compression fracture combined with intravertebral clefts. Intravertebral clefts have no significant impact on the effectiveness in the pain relief and function improvement.

The involvement of FoxO in cell survival and chemosensitivity mediated by Mirk/Dyrk1B in ovarian cancer.

Minibrain-related kinase (Mirk) is a serine/threonine kinase which is also known as the dual specificity tyrosine-phosphorylation-regulated kinase 1B (Dyrk1B). It is known that Dyrk1A, the closest family member to Mirk/Dyrk1B can mediate cellular localization of mammalian forkhead subclass O (FoxO1), a transcription factor, although the effect of Mirk/Dyrk1B on FoxO factors remains to be defined. In this study, we showed that Mirk/Dyrk1B protein was overexpressed in 5 of 8 ovarian cancer cell lines and negatively correlated with the protein level of FoxO factors (FoxO1+FoxO3A). Knockdown of Mirk by small interfering RNA (siRNA) resulted in cell apoptosis and sensitized cells to cisplatin accompanied by nuclear translocation of FoxO1 and/or FoxO3A as well as increased Bim, TRADD, cleaved caspase-3 and PARP. Furthermore, combined siRNAs of Mirk with FoxO1 and/or FoxO3A led to fewer apoptotic cells and cisplatin sensitivity compared to Mirk siRNA alone, suggesting that FoxO is involved in Mirk-mediated cell survival and chemosensitivity of ovarian cancer. Taken together, Mirk/Dyrk1B plays an important role in ovarian cancer cell survival through modulating FoxO translocation and may be a novel therapeutic target for ovarian cancer.

[Intervention assistant operation in treatment of phalanx closed fracture combined with artery crisis].

OBJECTIVE: To explore the effectiveness and methods of intervention assistant operation in the treatment of phalanx closed fracture combined with artery crisis. METHODS: Between August 2002 and December 2008, 24 cases (31 toes) of phalanx closed fracture combined with artery crisis were treated. There were 17 males (22 toes) and 7 females (9 toes), aged from 16 to 62 years (mean, 38 years). The causes of injury included crush and bruise (20 cases), traffic accident (3 cases), and machine twist (1 case). The locations were the first toe (19 toes), the second toe (10 toes), and the third toe (2 toes). The period between injury and hospitalization was 1-10 hours (mean, 6.8 hours). Phalanx angiography was performed by using venous indwelling needle for dorsalis pedis artery and posterior tibial artery puncture; according to angiography results, proper treatment could be done, then the contrast medium was injected to the artery to observe the blood supply. According to different types and locations of fracture, Kirschner wire and plate were chosen to fix fracture after the blood supply were recovered. RESULTS: Two cases (2 toes) received amputation due to necrosis at 4 days and 6 days after interventional therapy, respectively. Twenty-two cases (29 toes) survived. Incision healed primarily in 21 cases. Exudation occurred at wound of 1 case and was cured at 3 weeks after dressing change. Twenty-two cases (29 toes) were followed up 1-6 years (mean, 3.5 years) postoperatively. Two cases (3 toes) felt cool or anaesthesia and could not tolerate even in cold environment. The other toes had no senses of cold pain and paresthesia. Two cases (2 toes) had nonunion and achieved fracture healing after grafting bone. The mean union time was 4.5 months (range, 3-6 months) in other cases. CONCLUSION: Intervention assistant operation is an effective measure in the treatment of phalanx closed fracture combined with artery crisis.