FAR-RED ELONGATED HYPOCOTYLS3 negatively regulates shade avoidance responses in Arabidopsis.

Light is a key limiting factor of plant growth and development under the canopy. Specific light signals, such as a low ratio of red : far-red (R:FR) light, trigger the shade avoidance response, which affects hypocotyl, stem, and leaf growth. Although multiple components mediating shade avoidance responses have been identified in the past few decades, the underlying regulatory mechanism remains unclear. In this study, we found that the far-red elongated hypocotyls 3 (fhy3) mutant exhibited longer hypocotyls and increased expression levels of core shade avoidance response genes under low R:FR shade conditions compared with the wild type No-0, suggesting that FHY3 negatively regulates shade avoidance responses. Yeast one-hybrid, chromatin immunoprecipitation, and RT-qPCR assays revealed that FHY3 directly binds to the promoters and gene body of PHYTOCHROME RAPIDLY REGULATED1 (PAR1) and PAR2 and activates their expression to inhibit shade responses. Furthermore, the overexpression of PAR1 or PAR2 rescued the enhanced shade avoidance responses of fhy3, indicating that both genes are direct downstream targets of FHY3 that mediate shade avoidance responses. Our findings demonstrate that the light-signalling protein FHY3 positively regulates the transcription of PAR1 and PAR2, which encode two key negative regulators of shade avoidance responses, thus repressing plant responses to shade signals.

Effects of alfalfa flavonoids extract on the microbial flora of dairy cow rumen.

OBJECTIVE: The effect of flavonoids from alfalfa on the microbial flora was determined using molecular techniques of 16S ribosome deoxyribonucleic acid (rDNA) analysis. METHODS: Four primiparous Holstein heifers fitted with ruminal cannulas were used in a 4×4 Latin square design and fed a total mixed ration to which alfalfa flavonoids extract (AFE) was added at the rates of 0 (A, control), 20 (B), 60 (C), or 100 (D) mg per kg of heifer BW. RESULTS: The number of operational taxonomic units in heifers given higher levels of flavonoid extract (C and D) was higher than for the two other treatments. The Shannon, Ace, and Chao indices for treatment C were significantly higher than for the other treatments (p<0.05). The number of phyla and genera increased linearly with increasing dietary supplementation of AFE (p<0.05). The principal co-ordinates analysis plot showed substantial differences in the microbial flora for the four treatments. The microbial flora in treatment A was similar to that in B, C, and D were similar by the weighted analysis. The richness of Tenericutes at the phylum level tended to increase with increasing AFE (p = 0.10). The proportion of Euryarchaeota at the phylum level increased linearly, whereas the proportion of Fusobacteria decreased linearly with increasing AFE supplementation (p = 0.04). The percentage of Mogibacterium, Pyramidobacter, and Asteroleplasma at the genus level decreased linearly with increasing AFE (p<0.05). The abundance of Spirochaeta, Succinivibrio, and Suttonella at the genus level tended to decrease linearly with increasing AFE (0.05

Effects of alfalfa flavonoids on the production performance, immune system, and ruminal fermentation of dairy cows.

OBJECTIVE: The objective of this study was to examine the effects of alfalfa flavonoids on the production performance, immunity, and ruminal fermentation of dairy cows. METHODS: The experiments employed four primiparous Holstein cows fitted with ruminal cannulas, and used a 4×4 Latin square design. Cattle were fed total mixed ration supplemented with 0 (control group, Con), 20, 60, or 100 mg of alfalfa flavonoids extract (AFE) per kg of dairy cow body weight (BW). RESULTS: The feed intake of the group receiving 60 mg/kg BW of AFE were significantly higher (p<0.05) than that of the group receiving 100 mg/kg BW. Milk yields and the fat, protein and lactose of milk were unaffected by AFE, while the total solids content of milk reduced (p = 0.05) linearly as AFE supplementation was increased. The somatic cell count of milk in group receiving 60 mg/kg BW of AFE was significantly lower (p<0.05) than that of the control group. Apparent total-tract digestibility of neutral detergent fiber and crude protein showed a tendency to increase (0.05

Adult obese mice suffer from chronic secondary brain injury after mild TBI.

BACKGROUND: A traumatic brain injury (TBI) event is a devastating injury to the brain that may result in heightened inflammation, neurodegeneration, and subsequent cognitive and mood deficits. TBI victims with co-morbidities such as heart disease, diabetes, or obesity may be more vulnerable to the secondary brain injury that follows the initial insult. Compared to lean individuals, obese subjects tend to have worse clinical outcomes and higher mortality rates after trauma. METHODS: To elucidate whether obesity predisposes individuals to worse outcomes after TBI, we subjected adult lean and obese male/female mice to a mild TBI. The injury was administered using a controlled skull impact (CSI) device. Lean or obese 6-month-old C57 BL/6 mice were subjected once to a mild TBI. Additionally, at day 30 after injury, both the lean and obese mice were tested for increased anxiety using the open field test. RESULTS: At day 30 after TBI, compared to the lean mice, we found heightened microglial (MG) activation in the cerebral cortex, corpus callosum, and hypothalamus. Another compelling finding was that, compared to the non-injured obese male control mice, the obese TBI mice had a decrease in the rate of weight gain and serum corticosterone levels at day 30 after injury. Additionally, the injured obese mice displayed higher levels of anxiety as determined by a significant decrease in time spent in the non-peripheral zones in the open field test. In contrast to the obese males, the obese female mice did not exhibit increases in the number of active MG in the brain, changes in weight gain/corticosterone levels, or increased anxiety at day 30 after TBI. CONCLUSIONS: The data presented here suggests that obese mice have worse outcomes compared to lean mice after mild TBI. Also, the obese males have worse outcomes than the injured female mice. This data may explain the sequela of chronic secondary brain injury in obese adults after a single mild TBI. Also, this report may help shape how the overweight/obese populations are monitored over the days and months following a TBI.

Severe burn increased skeletal muscle loss in mdx mutant mice.

BACKGROUND: Severe burn causes muscle mass loss and atrophy. The balance between muscle cell death and growth maintains tissue homeostasis. We hypothesize that preexisting cellular structural defects will exacerbate skeletal muscle mass loss after burn. Using a Duchenne muscular dystrophy (mdx) mutant mouse, we investigated whether severe burn caused more damage in skeletal muscle with preexisting muscle disease. METHODS: The mdx mice and wild-type (WT) mice received 25% total body surface area scald burn. Gastrocnemius (GM), tibialis anterior, and gluteus muscles were obtained at days 1 and 3 after burn. GM muscle function was measured on day 3. Animals without burn served as controls. RESULTS: Wet tissue weight significantly decreased in tibialis anterior and gluteus in both mdx and WT mice after burn (P < 0.05). The ratio of muscle to body weight decreased in mdx mutant mice (P < 0.05) but not WT. Isometric force was significantly lower in mdx GM, and this difference persisted after burn (P < 0.05). Caspase-3 activity increased significantly after burn in both the groups, whereas HMGB1 expression was higher in burn mdx mice (P < 0.05). Proliferating cell nuclear antigen decreased significantly in mdx mice (P < 0.05). Myogenic markers pax7, myoD, and myogenin increased after burn in both the groups and were higher in mdx mice (P < 0.05). CONCLUSIONS: More muscle loss occurred in response to severe burn in mdx mutant mice. Cell turnover in mdx mice after burn is differed from WT. Although markers of myogenic activation are elevated in mdx mutant mice, the underlying muscle pathophysiology is less tolerant of traumatic injury.

Effects of exercise on soleus in severe burn and muscle disuse atrophy.

BACKGROUND: Muscle loss is a sequela of severe burn and critical illness with bed rest contributing significantly to atrophy. We hypothesize that exercise will mitigate muscle loss after burn with bed rest. MATERIALS AND METHODS: Male rats were assigned to sham ambulatory (S/A), burn ambulatory (B/A), sham hindlimb unloading (S/H), or burn hindlimb unloading (B/H). Rats received a 40% scald burn or sham and were ambulatory or placed in hindlimb unloading, a model of bed rest. Half from each group performed twice daily resistance climbing. Hindlimb isometric forces were measured on day 14. RESULTS: Soleus mass and muscle function were not affected by burn alone. Mass was significantly lower in hindlimb unloading (79 versus 139 mg, P < 0.001) and no exercise (103 versus 115 mg, P < 0.01). Exercise significantly increased soleus mass in B/H (86 versus 77 mg, P < 0.01). Hindlimb unloading significantly decreased muscle force in the twitch (12 versus 31 g, P < 0.001), tetanic (55 versus 148 g, P < 0.001), and specific tetanic measurements (12 versus 22 N/cm(2), P < 0.001). Effects of exercise on force depended on other factors. In B/H, exercise significantly increased twitch (14 versus 8 g, P < 0.05) and specific tetanic force (14 versus 7 N/cm(2), P < 0.01). Fatigue index was lower in ambulatory (55%) and exercise (52%) versus hindlimb (69%, P = 0.03) and no exercise (73%, P = 0.002). CONCLUSIONS: Hindlimb unloading is a significant factor in muscle atrophy. Exercise increased the soleus muscle mass, twitch, and specific force in this model. However, the fatigue index decreased with exercise in all groups. This suggests exercise contributes to functional muscle change in this model of disuse and critical illness.

[The effect of superior oblique tucking on the status of ocular torsion].

OBJECTIVE: To investigate the effect of superior oblique tucking on the status of ocular torsion. METHODS: It was a retrospective case series study. We reviewed the records of 25 patients underwent superior oblique tucking for treatment of superior oblique palsy in Tianjin Eye Hospital from January 2012 to and August 2013, among them, there were 12 patients had undergone inferior oblique myectomy or inferior rectus recession before this operation. All clinical records, including eye movement, vertical deviation, abnormal head position, Bielschowsky test et al were analyzed. Objective cyclotorsion were examined pre-operation, as well as 1, 30 days post-operation with fundus photograph. The photographs were transferred to a computer and then the fovea-disa angle (FDA) was measured by a software for drawing pictures. The difference of the two eyes was compared by the paired t-test, and the difference of the two groups was compared by the one-way analysis of variance. RESULTS: All patients had no more than 12 PD vertical deviation and significant abnormal head position before the operations. After underwent superior oblique tucking from 6 mm to 10 mm, their vertical deviation were less than 3 PD, and symptom of unacceptable abnormal head position were ameliorated or disappeared. In 25 cases of the monocular eye involvement, FDA was -10.60° ± 4.93° in the affected eyes preoperatively, while it was -9.67° ± 5.47° in the fellow eyes. There was also no statistically significant difference between two eyes (t = 0.54, P = 0.59). The combined FDA in both eyes were -20.27° ± 6.00° and -10.53° ± 8.04°, -10.83° ± 7.38° preoperatively, 1, and 30 days after operation. The comparison of objective ocular cyclotorsion for both eyes showed significant difference pre- and post-operation (F = 14.81, P < 0.01). CONCLUSIONS: Superior oblique tucking can correct ocular excyclodeviation effectively, and simultaneously correct little vertical deviation. It's good to ameliorate abnormal head position.

Discovery andw biological evaluation of novel 6,7-disubstituted-4-(2-fluorophenoxy)quinoline derivatives possessing 1,2,3-triazole-4-carboxamide moiety as c-Met kinase inhibitors.

A series of 6,7-disubstituted-4-(2-fluorophenoxy)quinoline derivatives possessing 1,2,3-triazole-4-carboxamide moiety were designed, synthesized and evaluated for their in vitro biological activities against c-Met kinase and five typical cancer cell lines (A549, H460, HT-29, MKN-45 and U87MG). Most compounds showed moderate to excellent antiproliferative activity. In this study, a promising compound 34, with a c-Met IC50 value of 1.04nM, was identified as a multitargeted receptor tyrosine kinase inhibitor. The SAR analyses indicated that compounds with halogen group, especially fluoro group, at 4-position on the phenyl ring (moiety B) have potent antitumor activity, and methylation on the 5-atom linker played an important role in the c-Met enzymatic activity.

Design, synthesis and pharmacological evaluation of 6,7-disubstituted-4-phenoxyquinoline derivatives as potential antitumor agents.

Two series of 6,7-disubstituted-4-phenoxyquinoline derivatives bearing 2,4-imidazolinedione/pyrazolone scaffold were designed, synthesized and evaluated for their c-Met kinase inhibition and cytotoxicity against HT-29, H460, A549, MKN-45, and U87MG cancer cell lines in vitro. The pharmacological data indicated that most of the tested compounds showed moderate to significant cytotoxicity and high selectivity against HT-29, H460 and A549 cancer cell lines as compared with foretinib. The SAR analyses indicated that compounds with halogen groups, especially trifluoromethyl groups at 2-position on the phenyl ring (moiety B) were more effective. In this study, a promising compound 17 (c-Met IC50=2.20nM, a multi-target tyrosine kinase inhibitor) showed the most potent antitumor activities with IC50 values of 0.14µM, 0.18µM, 0.09µM, 0.03µM, and 1.06µM against HT-29, H460, A549, MKN-45, and U87MG cell lines, respectively.

[The effect of superior oblique recession on the status of ocular torsion].

OBJECTIVE: To investigate the effect of superior oblique recession on the status of ocular torsion. METHODS: Twenty patients (31 eyes ) underwent superior oblique recession for treatment of superior oblique overacting, 11 cases had both eyes involvement while 9 had monocular.Objective cyclotorsion were examined pre-operation, as well as 1, 30, 90 days post-operation with fundus photograph. The photographs were transferred to a computer and then the fovea-disa angle (FDA) was measured by a software for drawing pictures.Subjective cyclotorsion were assessed by double Maddox rod test and synoptophore, while the binocular vision function by Bagolini glasses and Titmus. RESULTS: For 11 cases of the both eyes involvement, preoperation FDA was 9.50° ± 5.12° in the right eyes, 8.23° ± 3.45° in the left eyes, there was not statistically significant between two eyes(t = 1.12, P = 0.29). For 9 cases of the monocular eye involvement, preoperation FDA was 6.55° ± 4.14° in the affected eyes, 6.85° ± 5.08° in the follow eyes, there was also not statistically significant between two eyes (t = 0.13, P = 0.90). To 11 cases of the both eyes involvement, the combined FDA of preoperation and 1, 30, 90 days after operation were 17.74° ± 7.89° and 7.63° ± 4.28°, 6.05° ± 6.11°, 7.33° ± 6.17° respectively. The comparision of objective ocular cyclotorsion for both eyes showed significant difference pre-and post-operation (F = 8.245, P < 0.01) . To 9 cases of the monocular eye involvement, the combined FDA of preoperation and 1, 30, 90 days after operation were 13.40° ± 6.13° and 5.06° ± 3.99°, 3.59° ± 3.47°, 5.07° ± 3.35° respectively. The comparision of objective ocular cyclotorsion showed significant difference pre-and post-operation (F = 9.351, P < 0.01) .Four patients had subjective cyclotorsion, but it's different with objective cyclotorsion before and after surgery.8 patients' binocular vision function were ameliorated after surgery. CONCLUSION: Recession of overaction superior oblique muscles can correct ocular incyclodeviation effectively.

Identification of the forkhead transcriptional factor 2 (FOXL2) gene mutations in four Chinese families with blepharophimosis syndrome.

PURPOSE: To determine the genetic origin of disease in four Chinese families with blepharophimosis syndrome. METHODS: Four Han Chinese families with blepharophimosis syndrome were ascertained and patients underwent complete physical and ophthalmic examinations. Blood samples were collected and genomic DNA was extracted. Sequence analysis of the forkhead transcriptional factor 2 (FOXL2) gene was performed by direct sequencing and mutations were analyzed. RESULTS: Three mutations in FOXL2 were found in four families, including c.672_701dup30 (p.Ala224_Ala234dup10), c.313C>A (p.N105H), and c.430G>T (p.R144W). The c.672_701dup30 (p.Ala224_Ala234dup10) mutation was reported previously and predicted to result in expansions of the polyalanine tract. The mutations of c.313C>A (p. N105H) and c.430G>T (p.R144W) are two novel missense mutations. CONCLUSIONS: Our study further supports the view that the expansion of the polyalanine tract is the hotspot of mutations within FOXL2. The two novel missense mutations detected in this study will expand the mutation spectrum of the FOXL2 gene and contribute to the research on the molecular pathogenesis of FOXL2.

[Recession of both horizontal rectus muscles in Duane Retraction Syndrome with significant globe retraction].

OBJECTIVE: To evaluate the effect of Recession of both horizontal rectus muscles in Duane Retraction Syndrome with significant globe retraction. METHODS: Sixteen cases with DRS were summarized retrospectively. All patients had undergone surgery with recession of both horizontal rectus muscles. All clinical records, including sex, age, types of DRS, clinical features, surgical methods and clinical outcomes were analyzed. All patients were followed up for 3 months to 1 year. RESULTS: Fifteen cases had only monocular involvement while one had both eyes. The number of type I DRS was 3 cases, 1 case was esotropia while others were orthotropic in primary position. Type III DRS was observed 13 cases. Esotropia was seen in 6 cases (7 eyes), exotropia of 1 cases and orthotropic in primary position of 6 cases. 10 cases exhibited marked face turn. An upshoot or downshoot and variable severity of retraction of globe were found in all patients on attempt adduction of the affected eye. All patients had undergone surgery with recession of both horizontal rectus muscles. The medical rectus muscles were recessed from 5 mm to 7 mm and lateral rectus muscles 3 mm to 9 mm simultaneously, which was based on the amount of primary position deviation. Among these 2 cases were combined with Y-splitting of lateral rectus muscle. After surgery, all patients were orthotropic in primary position. Their symptom of unacceptable abnormal head position, significant globe retraction, noticeable narrowing of the palpebral fissure and significant upshoot or downshoot were ameliorated or disappeared. Especially the recession of lateral rectus muscle in addition to Y-splitting combining with the simultaneous medial rectus recession resulted in further amelioration of globe retraction in addition to upshoot and downshoot. CONCLUSION: Recession of both horizontal rectus muscles is effective in the treatment of significant globe retraction in Duane syndrome. Type III DRS with significant globe retraction but has no marked deviation and face turn can adopt this method to ameliorate their aspect. The method of lateral rectus muscle in addition to Y-splitting plays an important role in the treatment of upshoot and downshoot.

The cGAS-STING pathway connects mitochondrial damage to inflammation in burn-induced acute lung injury in rat.

OBJECTIVE: Damage associated molecular patterns (DAMPs) are pathological mediators linking local tissue damage to systemic inflammation in various diseases. Some DAMPs, such as mitochondrial DNA (mtDNA), can be recognized by the cytoplasmic cGAS protein to trigger the activation of the stimulator of interferon genes (STING)-dependent innate immune pathway responsible for infection or sterile inflammation. The objective of our study was to evaluate the association between circulating mtDNA and cGAS-STING pathway activation in mediating inflammation following burn injury. METHODS: 48 adult Sprague-Dawley male rats were divided into eight groups (Sham, 2, 4, 8, 12, 24, 48, 72 h after burn injury). The animals underwent 40% total body surface area scald injury to produce a full-thickness burn. Plasma samples were collected via cardiac puncture under deep anesthesia. Tissues were harvested and placed in formalin, followed by paraffin embedment. Total plasma DNA was isolated followed by measurement of mtDNA using quantitative polymerase chain reaction. Haemotoxylin-Eosin stain and Western blot was used for lung histology and protein assays, respectively. Statistical analyses were performed using ANOVA and student's t-test and represented as mean ± s.d. RESULTS: Plasma mtDNA trended upward at early time-points following burn injury with peak levels at 8 h after burn when compared to the control group (345 ± 83.4 copies/µl vs. 239 ± 43.1 copies/µl, p = 0.07) and followed a bell-shaped distribution. Lung slices from burned rats showed acute injury marked by increased inflammatory infiltrate, with the maximum changes seen at 24 h, accompanied with significant upregulation of neutrophil elastase (p = 0.04). Compared with sham animals, cGAS and STING protein levels in lung tissue were up-regulated at 4 and 8 h after burn (p = 0.03 and p < 0.001, respectively). CONCLUSION: Activation of the cGAS-STING pathway by increased plasma mtDNA is an important pathway driving neutrophil infiltration in burn-induced acute lung injury in rats. A further understanding of the STING-mediated immunopathology in lung and other susceptible organs may be important for the development of novel therapies for burn injury.

IL-10 polymorphism associated with decreased risk for mortality after burn injury.

OBJECTIVE: Evaluation of single nucleotide polymorphisms (SNPs) in the interleukin-10 promoter (-592 and -819) on risk for death after burn injury. METHODS: Association between the IL-10 SNPs and outcome after burn injury was evaluated in a cohort of 265 patients from Parkland Hospital, Dallas, TX with ≥ 15% TBSA burns without non-burn trauma (ISS ≤ 16), traumatic or anoxic brain injury or spinal cord injury, who survived >48 h under an IRB-approved protocol. Clinical data were collected prospectively and genotyping was conducted by TaqMan assay. Whole blood from 31 healthy volunteers was stimulated with LPS (100 ng/mL) to determine the level of IL-10 expression for each allele by enzyme-linked immunosorbent assay (ELISA). RESULTS: After adjustment for percent total body surface area (TBSA) burned, inhalation injury, age, gender, and race/ethnicity, carriage of ­592A and/or ­819T was significantly associated (P = 0.014) with a decreased risk for death (adjusted odds ratio: 0.404; 95% CI: 0.197-0.829). As the candidate SNPs were in complete linkage disequilibrium, it was not possible to distinguish which allele was associated with decreased mortality risk. Age, inhalation injury, and full-thickness burn size were significantly associated with increased risk for death. In the LPS stimulated blood of healthy controls, carriage of the -592A and/or -819T allele demonstrated a trend for decreased levels of IL-10 (P = 0.079). CONCLUSION: Carriage of the ­592A and/or ­819T allele in the IL-10 promoter appears to reduce the risk for death after burn injury.

Tau Is Elevated in Pediatric Patients on Extracorporeal Membrane Oxygenation.

Neurologic injury is a known and feared complication of extracorporeal membrane oxygenation (ECMO). Neurologic biomarkers may have a role in assisting in early identification of such. Axonal biomarker tau has not been investigated in the pediatric ECMO population. The objective of this study is to evaluate plasma levels of tau in pediatric patients supported with ECMO. Eighteen patients requiring ECMO support in a quaternary pediatric intensive care unit at a university-affiliated children's hospital from October 2015 to February 2017 were enrolled. Patients undergoing extracorporeal cardiopulmonary resuscitation or recent history of bypass were excluded. Plasma tau was measured using enzyme-linked immunosorbent assay. Neuroimaging was reviewed for acute neurologic injury, and tau levels were analyzed to assess for correlation. Tau was significantly higher in ECMO patients than in control subjects. Sixty-one percent of subjects had evidence of acute brain injury on neuroimaging, but tau level did not correlate with injury. Subjects with multifocal injury all experienced infarction and had significantly higher tau levels on ECMO day 3 than patients with isolated injury. In addition, peak tau levels of neuro-injured subjects were compared with controls and noninjured ECMO subjects using receiver operating curve analysis. This study demonstrates preliminary evidence of axonal injury in pediatric ECMO patients.

Structure of a green algal photosystem I in complex with a large number of light-harvesting complex I subunits.

Photosystem I (PSI) is a highly efficient natural light-energy converter, and has diverse light-harvesting antennas associated with its core in different photosynthetic organisms. In green algae, an extremely large light-harvesting complex I (LHCI) captures and transfers energy to the PSI core. Here, we report the structure of PSI-LHCI from a green alga Bryopsis corticulans at 3.49 Å resolution, obtained by single-particle cryo-electron microscopy, which revealed 13 core subunits including subunits characteristic of both prokaryotes and eukaryotes, and 10 light-harvesting complex a (Lhca) antennas that form a double semi-ring and an additional Lhca dimer, including a novel four-transmembrane-helix Lhca. In total, 244 chlorophylls were identified, some of which were located at key positions for the fast energy transfer. These results provide a firm structural basis for unravelling the mechanisms of light-energy harvesting, transfer and quenching in the green algal PSI-LHCI, and important clues as to how PSI-LHCI has changed during evolution.

Correlation analysis on the expression levels of microRNA-23a and microRNA-23b and the incidence and prognosis of ovarian cancer.

The present study aimed to investigate the expression levels of microRNA (miR)-23a and miR-23b in the tumor tissues of patients with ovarian cancer. The study also explored the correlation of miR-23a and miR-23b expression levels in the tumor tissues with the clinic-pathological parameters and prognosis of the patients. Specimens of frozen tumor tissues and normal tissues adjacent to the tumor were collected from 50 patients with ovarian cancer. Reverse transcription-quantitative polymerase chain reaction was adopted to detect the expression levels of miR-23a and miR-23b in tumor tissues. Furthermore, normal tissues adjacent to the tumor were utilized as the control for the experiments and Pearson method was used to analyze the correlation between miR-23a and miR-23b expression levels in tumor tissues. The correlation of miR-23a and miR-23 expression in tumor tissues and the prognosis of the patients with ovarian cancer was analyzed in combination with clinical data. The expression of miR-23a in the tissues of ovarian cancer was significantly higher in comparison with normal adjacent tissues. However, the expression of miR-23b in the tissues of ovarian cancer was significantly lower when compared with adjacent normal tissues. Notably, the expression of miR-23a was negatively correlated with that of miR-23b in tumor tissues of ovarian cancer. The high expression of miR-23a and the low expression of miR-23b in tumor tissues of the patients was correlated with the degree of tumor differentiation, metastasis of lymph nodes and clinical staging. The five-year overall survival rate of the patients was 36% (18/50). Univariate survival analysis indicated that miR-23a and miR-23b were the factors influencing the overall survival rate of ovarian cancer. The present findings suggest that high expression of miR-23a and the low expression of miR-23b are closely correlated with the occurrence and development of ovarian cancer. The abnormal expression of the miR-23a and miR-23b could be utilized as potential prognostic molecular markers of ovarian cancer.

Optimization of low-abundance protein extraction and abundant protein removal from defatted soybean meal.

The aim of this study was to optimize the conditions for the extraction of low-abundance proteins (LAPs) and the removal of abundant proteins (APs; ß-conglycinin and glycinin) from soybean meal. Single factor and orthogonal experiments were designed to determine the effects of four factors (isopropanol concentration, total extraction time, ultrasonic power, and ultrasonic time) on protein concentration in isopropanol extracts. Proteins in the isopropanol supernatant and the cold acetone precipitate of isopropanol were identified by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF-MS). The results showed that the optimal conditions were 50% isopropanol, ultrasonic pretreatment for 15 min at 350 W, and a total extraction time of 1 h. Under these conditions, the protein concentration in the isopropanol extracts reached 0.8081 g/L. Many LAPs were detected, including ß-amylase, soybean agglutinin, soybean trypsin inhibitor, fumarylacetoacetase-like, phospholipase D alpha 1-like, oleosin, and even some unknown soybean proteins. The soybean APs (ß-conglycinin and glycinin) were not found. The method may be useful for discovering new soybean proteins and extracting enough LAPs of soybean to allow further studies of their physiological effects on animals without the influence of APs.

Establishment of primary bovine intestinal epithelial cell culture and clone method.

The aim of this study was to establish bovine intestinal epithelial cell (BIEC) line and provide a novel clone cell method. Although various strategies of bovine cell culture and clone techniques have been reported, these methods remain not established. Here, we culture successfully primary BIECs and establish a novel clone cell method. Our result showed that BIECs could be successfully cultured and passaged about generation 5. These cellular aggregates and clusters were adherent loosely at day 2 of culture. Cell aggregates and clusters start to proliferate after approximately 4 d. The BIECs showed positive reaction against cytokeratin 18, E-cadherin, and characteristics of epithelial-like morphology. In addition, the fatty acid-binding proteins (FABPs), villin, and intestinal peptidase (IP) band were positive in BIECs. Our results suggest that the establishment of culturing and clone BIEC methods will apply to isolate and clone other primary cells. These BIECs could therefore contribute to the study of bovine intestinal nutrient absorption and regulation, immune regulation, and the pathogenesis of the bovine intestinal disease, which will provide intestinal cell model in vitro.

Three-dimensional culture system can induce expression of casein in immortalized bovine mammary epithelial cells.

Primary bovine mammary epithelial cells (BMECs) are not ideal models for long-term studies of lactation mechanisms because these cells in a monolayer culture system cannot be polarized to simulate the physiological functions in vitro. We investigate the effects of different culture models and karyotypes on casein expression in a three-dimensional (3D) culture system. The immortalized cells' karyotypes were analyzed at passages 10, 20, 30 and 40 to detect the effects of chromosome stability. Western blotting examined that whether or not the immortalized cells at passages 5, 10, 20, 30, 40 and 50 could induce expression of casein in a 3D culture system. The proper polarization of the acinar structures was monitored. BMECs were successfully immortalized. The cell karyotype at passage 30 remained at 60 chromosomes and the average value was 57.1 ± 0.40 after passage 40. The polarized protein's levels were up-regulated in 3D culture compared to 2D culture. Expression of αs1, ß and κ-casein could be detectable in a passage range in 3D culture. Expression of αs2-casein was undetectable in all experimental groups. However, all casein expressions were barely detectable in traditional 2D culture system. Therefore, 3D culture system is an important tool for the long-term study of lactation mechanisms in vitro.