[Research progress on exosomes and their non-coding RNAs in the diagnosis of neurodegenerative diseases].

Neurodegenerative diseases, originating from irreversible progressive loss of neuronal structure or function, are difficult to diagnose and treat. They vary widely in scope and have poor prevention and prognosis. Therefore, research on their early diagnosis is particularly important. Exosomes are small vesicles of cellular origin that contain various bioactive small molecules, such as proteins, RNAs, and DNAs, and play important roles in intercellular communication. Recent studies have shown that exosomes and their non-coding RNAs are key factors in the pathogenesis of various neurodegenerative diseases. Therefore, exosomes and their non-coding RNAs may provide a breakthrough for the early diagnosis of neurodegenerative diseases. This review summarizes the biology of exosomes and the current research progress of exosomes and their non-coding RNAs in diagnosing neurodegenerative diseases and further explores the challenges and prospects they face.

[The prevention and therapy of corneal ulcers in patients with large-area thermal burns].

Objective: To evaluate the characteristics of ocular injury in patients with severe extensive thermal burns, and to explore the effective methods to prevent and treat corneal ulcers related to severe burns. Methods: A retrospective case series study. Between 2010 and 2019, Sixteen severe thermal burn patients with burn sizes>70% of the total body surface area involving the ocular region were admitted to the Burns and Plastic Surgery Division of Chinese PLA General Hospital, and consult with Ophthalmology Division. There were deep second-degree to fourth-degree burns in the eyelids. In the eleven surviving patients, 22 eyes presented ectropion. Eyelid full-thickness skin grafting (EFTSG) combined with or without tarsorrhaphy was performed in 20 eyes due to severe corneal exposure. Two eyes received partial blepharorrhaphy because of mild ectropion. The ocular manifestations and treatment outcomes were reviewed and assessed. Results: The majority of the patients were youth, and the average age was (36.8±10.4) years. The burn area was 84.0%±9.1% of the body surface area. Corneal ulcers secondary to lagophthalmos occurred at (35.1±15.6) days after burning in 75% (24/32) of eyes. Perforation was found in 18.8% (6/32) of eyes. Among the 22 operated eyes, the corneal ulcer was repaired in all 9 eyes receiving EFTSG with tarsorrhaphy, whereas ectropion recurred in 8 of 11 eyes only receiving EFTSG, and 4 eyes underwent further surgery due to corneal epithelial defects. Conclusions: In patients with severe large-area thermal burns, corneal ulcers are common complications. Prevention of corneal exposure is vital because the treatment of corneal ulceration is difficult due to eyelid deformity, inflammation and the absence of donor skin. Timely full-thickness skin grafting and blepharorrhaphy are effective approaches to preventing exposure keratopathy. To severe ulcers occur, conjunctival flap or Tenon's capsule covering combined with eyelid EFTSG and tarsorrhaphy is useful to rescue visual function.

Enhanced removal of Congo red dye from aqueous solution by surface modified activated carbon with bacteria.

AIMS: The adsorption behaviour and mechanisms of the surface modified activated carbon with bacteria was evaluated. METHODS AND RESULTS: 16S rRNA was employed to identify the hydrocarbon-degrading bacteria. The bacteria was characterized by TEM and electron microscope. The surface modified activated carbon with bacteria was characterized by SEM. The adsorption behaviour was tested by static adsorption and dynamic adsorption. CONCLUSION: The adsorption efficiency of the modified activated carbon was high when pH was weak acidic, and the adsorption capacity increased with the increase of temperature ranging from 20 to 35°C. The adsorption capacity peaked at 234·6 mg g-1 at 25°C, which was sixfold higher than that of activated carbon. The pseudo-first-order kinetic can more accurately assess Congo red adsorption on the two adsorbents. The adsorption of Congo red by bacteria surface modified activated carbon fitted well with the Langmuir's model. The adsorption process was endothermic, and the biological floccules were formed during the adsorption. The physical adsorption is the main driving force. SIGNIFICANCE AND IMPACT OF THE STUDY: The results indicate that the bacteria surface-modified activated carbon can be used effectively as an adsorbent to eliminate Congo red from aqueous solutions.

[Characteristics and related factors of plantar pressure in the chronic ankle instability individuals].

OBJECTIVE: To analyze characteristics and related factors of the plantar pressure during the level walking and single leg standing in the chronic ankle instability (CAI) individuals. METHODS: From April 2019, 75 CAI individuals and 40 healthy individuals were enrolled in this study. Both of the static and dynamic plantar pressure were measured during six times level walking and three times single leg standing testing. The data including peak force, time to peak force in various foot contact areas and the time to boundary (TTB) and velocity of center of pressure (COP) were measured and compared between the affected side and the unaffected side and between the CAI cases and the healthy individuals. The correlations between the plantar pressure and the gender, Beighton score, affected side and body mass index (BMI) were analyzed. RESULTS: The characteristics of plantar pressure distribution in the CAI individuals included: (1) During the level walking, the affected side showed the similar pressure contribution as the unaffected side (P>0.05). While compared with healthy individuals, there was a significantly higher peak force in the 5th metatarsal area (t=-3.86, P=0.03) of the affected side, lower peak force in the 1st (t=2.99, P=0.02), 2nd metatarsal head areas (t=2.09, P=0.01) of the affected side, medial hindfoot areas of both sides (affected, t=2.33, P=0.01; unaffected, t=3.74, P=0.02) and toes areass of both sides (affected, t=2.23, P=0.01; unaffected, t=3.28, P=0.02) and a delay to peak force in the 4th metatarsal head area (t=3.33, P=0.01) of the affected side. (2) During the single leg standing, the CAI individuals showed significantly worse balance control in the anterior/posterior direction (P < 0.05) and lateral/medial direction (P < 0.05) compared with the healthy controls, and the affected side had more severe balance control deficit in the lateral/medial direction (P < 0.05). (3) The women (P < 0.05) and the individuals with higher Beighton scores (P < 0.05) showed worse balance control deficit in the lateral/medial direction. CONCLUSION: CAI individuals showed significantly a more lateral shifted plantar distribution during the level walking compared with the healthy individuals and the tendency was worse on the affected sides, and showed worse balance control in the anterior/posterior direction and lateral/medial direction during the single leg standing. The women and those with generalized ligament laxity showed significantly worse balance control.

[Complexity of Detecting CRISPR/Cas9-Mediated Homologous Recombination in Zebrafish].

Homology-directed (HD) genome modification offers an opportunity to precisely modify the genome. Despite reported successful cases, for many loci, precise genome editing remains challenging and inefficient in vivo. Here we report an effort to precisely knock-in a GFP reporter into gad locus mediated by CRISPR/Cas9 system in the zebrafish Danio rerio. PCR artifact was detected in testing for homologous recombination (HR), but was mitigated by optimizing PCR condition and decreasing the injected targeting plasmid concentration. Under this optimized condition, time course analysis revealed a decline of the HR-positive embryos at embryogenesis progressed. GFP signals also diminished at later developmental stages. The GFP signals were consistent with PCR detection, both of which suggested the loss of targeted insertion events at later stages. Such loss of insertion might be one underlying reason for the inability to obtain germ-line transgenic lines with GFP knocked into the gad locus. Our results suggest that the low HR efficiency associated with CRISPR-mediated knock-in is in part due to loss of insertion after targeted integration into the gad locus.

[Mid-to-long term outcomes and influence factors of postoperative concurrent chronic ankle instability and posterior ankle impingement].

OBJECTIVE: To investigate the mid-to-long-term efficacy of patients with chronic ankle instability combined with posterior impingement syndrome after 3-9 years of follow-up, and to analyze the influencing factors. METHODS: From February 2010 to December 2015, 46 patients underwent concurrent lateral ankle ligament repair with posterior ankle arthroscopic surgery at the Institute of Sports Medicine, Peking University Third Hospital. The patient was first placed in a prone position and underwent arthroscopic debridement for the posterior impingement. After finishing the posterior arthroscopy, the surgeon and assistants first translated the patient to the affected side, then turned to the healthy side, and changed the position to the supine position. During the turning over, another assistant held the arthroscope and the instrument to ensure that it was sterile and could be used without replacement. The anterior ankle arthroscopy was operated if necessary and the lateral ankle ligament repair was anatomic repaired with anchors. The 42 patients were followed up, including 26 males and 16 females. The average age was (28.9±10.0) years. The patient's clinical symptoms, joint stability, mobility and motor function were compared by questionnaire and physical examination. The preoperative and postoperative visual analogue scale (VAS) scores, American Orthopaedic Foot and Ankle Society (AOFAS) scores, Tegner scores were compared, and the clinical scores and the patient age, gender, height and weight were compared. The correlations between body mass index (BMI), preoperative duration, surgery duration, and cartilage injury were analyzed. RESULTS: The mean follow-up time was (71.8±22.8) months. The postoperative VAS scores (1.0 vs. 5.0, P<0.001), AOFAS scores (92 vs. 80, P<0.001), and Tegner scores (6.5 vs. 2, P<0.001) were significantly superior to the preoperative levels. The excellent and good rate was 97.6%. The postoperative VAS score (t=2.719, P=0.10), AOFAS score (t=-2.853, P=0.10), Tegner score (t=-3.443, P=0.001) and time to return exercise (t=2.814, P=0.008) were negatively correlated with the patient age, and the postoperative VAS score was negatively correlated with cartilage injury (Z=-2.195, P=0.028). CONCLUSION: The mid-to-long-term clinical outcomes of the chronic ankle ligament instability combined with the posterior impingement were good. The age of the patients was negatively correlated with the clinical outcome. The combined cartilage injury could aggravate the postoperative pain.

Sex-related differences in response to zinc pyrithione shampoo vs. non-anti-dandruff shampoo.

OBJECTIVE: Sex-related differences in skin properties may be expected to impact dandruff formation and treatment. A meta-analysis approach was undertaken to investigate potential differences between males and females in response to zinc pyrithione (ZnPT) treatment vs. non-anti-dandruff (AD) shampoo. A separate pooled statistical analysis of ceramide and total protein loss endpoints was also undertaken to assess potential sex-related differences in stratum corneum properties that might influence response to ZnPT vs. non-AD shampoo in subjects with dandruff. METHODS: The meta-analysis approach included data from 17 half-head, double-blind, randomized studies (N = 2088) undertaken in Asia to assess the effectiveness of 1% ZnPT shampoo and/or non-AD shampoo in reducing dandruff severity, as assessed by Total Weighted Head Score Adherent Flake (TWHS-AF) methodology. Treatment duration was 4 weeks, with TWHS-AF measured at weekly intervals. Data from an additional three studies (N = 143) conducted in Asia were included in the pooled analysis of ceramide levels and protein loss from scalp skin of subjects with dandruff. RESULTS: Response to 1% ZnPT vs. non-AD shampoo was greater in males than in females at all time points; after 4 weeks, the between-treatment difference in TWHS-AF was -17.5 (95% confidence interval [CI] -19.5, -15.5) in males and -11.1 (-13.2, -8.9) in females. Sex-related differences were observed between males and females in response to both 1% ZnPT and non-AD shampoos. Males had a stronger response than females to treatment with 1% ZnPT shampoo, while dandruff decreased to a greater extent in females than in males when using non-AD shampoo. Statistically significant sex-related differences in ceramide levels and total protein loss were observed (both P < 0.01). Ceramide levels were 0.76 times lower (95% CI 0.60, 0.97) in males than in females, while total protein loss was 1.4 times greater (95% CI 1.1, 1.9) in males than in females. CONCLUSION: Males show a greater response than females to 1% ZnPT shampoo, while females show a greater response than males to non-AD shampoo. These findings may in part be explained by the sex-related differences observed in stratum corneum properties, which may make males more prone to dandruff than females.

Effectiveness of rubber seed oil and flaxseed oil to enhance the α-linolenic acid content in milk from dairy cows.

This experiment was conducted to investigate effect of rubber seed oil compared with flaxseed oil when fed alone or in combination on milk yield, milk composition, and α-linolenic acid (ALA) concentration in milk of dairy cows. Forty-eight mid-lactation Holstein dairy cows were randomly assigned to 1 of 4 treatments according to a completely randomized design. Cows were fed a basal diet (control; CON) or a basal diet supplemented with 4% rubber seed oil (RO), 4% flaxseed oil (FO), or 2% rubber seed oil plus 2% flaxseed oil (RFO) on a dry matter basis for 9 wk. Feed intake, milk protein percentage, and milk fat levels did not differ between the treatments. Cows fed the RO, FO, or RFO treatments had a higher milk yield than the CON group (up to 10.5% more), whereas milk fat percentages decreased. Compared with the CON, milk concentration of ALA was substantially higher in cows receiving RO or RFO, and was doubled in cows receiving FO. The ALA yield (g/d) increased by 31.0, 70.3, and 33.4% in milk from cows fed RO, FO, or RFO, respectively, compared with the CON. Both C18:1 trans-11 (vaccenic acid) and C18:2 cis-9,trans-11 (conjugated linoleic acid; CLA) levels were higher in cows fed added flaxseed or rubber seed oil. The CLA yield (g/d) increased by 336, 492, and 484% in cows fed RO, FO, or RFO, respectively, compared with the CON. The increase in vaccenic acid, ALA, and CLA was greater in cows fed RFO than in cows fed RO alone. Compared with the CON, the milk fat from cows fed any of the dietary supplements had a higher concentration of unsaturated fatty acids, monounsaturated fatty acids, and polyunsaturated fatty acids; conversely, the saturated fatty acids levels in milk fat were 30.5% lower. Insulin and growth hormones were not affected by dietary treatments; however, we noted an increase in both cholesterol and nonesterified fatty acids levels in the RO, FO, or RFO treatments. These results indicate that rubber seed oil and flaxseed oil will increase milk production and the concentration of functional fatty acids (ALA, vaccenic acid, and CLA) in milk fat while decreasing the content of saturated fatty acids.

Intra-articular delivery of anti-Hif-2α siRNA by chondrocyte-homing nanoparticles to prevent cartilage degeneration in arthritic mice.

Hypoxia-inducible factor-2α (Hif-2α) is a potential therapeutic target for osteoarthritis (OA), but the application of this target in the delivery of therapeutic agents to chondrocytes remains a challenge. A chondrocyte-targeting vector was constructed in a previous study to enhance transfection efficiency and specificity of chondrocytes in vivo. This study used vectors to deliver small-interfering RNA (siRNA) and silenced Hif-2α expression to prevent cartilage degeneration in OA-affected mice. After siRNA transfection was conducted by cartilage-targeting nanoparticles, the protein levels of Hif-2α, matrix metalloproteinases (MMP-13, -9), a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS-4, -5), vascular endothelial growth factor (VEGF), type X collagen and nuclear factor (NF)-κB in interleukin-1-beta (IL-1ß)-stimulated chondrocytes were determined. Chondrocyte-targeting ability was also determined by fluorescein isothiocyanate (FITC)-labeled siRNA tracking under a confocal microscope. OA model was established by surgically destabilizing the knee joints of a mouse. Hif-2α siRNA was then delivered intra-articularly with nanoparticles in vivo. Cartilage degeneration and synovium inflammation in the knee joints were analyzed by histomorphometry. IL-1ß levels in the synovial fluid were also measured by enzyme-linked immunosorbent assay. In vitro assay results showed that catabolic factors, including Hif-2α, MMP-13 and -9, ADAMTS-4, VEGF, collagen type X and NF-κB, were downregulated after Hif-2α-siRNA transfection by chondrocyte-targeting nanoparticles. In vivo assay results with FITC-labeled siRNA tracking also confirmed that nanoparticles promoted the local concentration and prolonged the retention time of siRNA in the cartilage. Histological analysis results confirmed that nanoparticle-mediated siRNA maintained cartilage integrity and alleviated synovium inflammation. IL-1ß levels decreased after siRNA was silenced by nanoparticles. Thus, chondrocyte-targeting nanoparticles could deliver Hif-2α siRNA to cartilage and specifically inhibit the expression of catabolic proteins.

Cognitive motor interference for gait and balance in stroke: a systematic review and meta-analysis.

BACKGROUND AND PURPOSE: An increasing interest in the potential benefits of cognitive motor interference (CMI) for stroke has recently been observed, but the efficacy of CMI for gait and balance is controversial. A systematic review and meta-analysis of randomized controlled trials was performed to estimate the effect of CMI on gait and balance in patients with stroke. METHODS: Articles in Medline, EMBASE, the Cochrane Library, Web of Science, CINAHL, PEDro and the China Biology Medicine disc were searched from 1970 to July 2014. Only randomized controlled trials examining the effects of CMI for patients with stroke were included, and no language restrictions were applied. Main outcome measures included gait and balance function. RESULTS: A total of 15 studies composed of 395 participants met the inclusion criteria, and 13 studies of 363 participants were used as data sources for the meta-analysis. Pooling revealed that CMI was superior to the control group for gait speed [mean difference (MD) 0.19 m/s, 95% confidence interval (CI) (0.06, 0.31), P = 0.003], stride length [MD 12.53 cm, 95% CI (4.07, 20.99), P = 0.004], cadence [MD 10.44 steps/min, 95% CI (4.17, 16.71), P = 0.001], centre of pressure sway area [MD -1.05, 95% CI (-1.85, -0.26), P = 0.01] and Berg balance scale [MD 2.87, 95% CI (0.54, 5.21), P = 0.02] in the short term. CONCLUSION: Cognitive motor interference is effective for improving gait and balance function for stroke in the short term. However, only little evidence supports assumptions regarding CMI's long-term benefits.

Detection of the genetic variation of polygalacturonase-inhibiting protein gene 2 in autotetraploid alfalfa (Medicago sativa) using an improved SSCP technique.

In this study, 2 approaches were adopted to obtain good single-strand conformation polymorphism (SSCP) data for autotetraploid alfalfa; primers were added to PCR products, and fluorescent-labeled primers were utilized. PCR-SSCP conditions for a 331-bp fragment in the coding region of polygalacturonase-inhibiting protein gene 2 in alfalfa (MsPGIP2) were optimized, and the results showed that the best SSCP gel pattern could be obtained when the loading mixture was made by mixing 1 µL PCR products, 0.2 to 0.8 µL unlabeled primers (50 µM) and 4 to 16 µL loading buffer. Furthermore, the use of the fluorescent-labeled primers resulted in 2 separated electrophoresis images from 2 complementary single DNA strands, thus making the determination of alleles and idiotypes a relatively easy task. In addition, the results of sequencing prove that the determination of alleles and idiotypes were accurate based on SSCP analysis. Finally, a total of 9 alleles with 18 SNP sites were identified for MsPGIP2 in the alfalfa variety 'Algonquin'. In conclusion, MsPGIP2 possessed great genetic variation, and the addition of primers to the PCR products in combination with the fluorescent labeling of primers could significantly improve the sensitivity and resolution of SSCP analysis. This technique could be used for genetic diversity detection and marker-assisted breeding of useful genes in autopolyploid species such as alfalfa.

Significance of Bcl10 gene mutations in the clinical diagnosis of MALT-type ocular adnexal lymphoma in the Chinese population.

We investigated the expression of Bcl10 gene mutations in mucosa-associated lymphoid tissue-type ocular adnexal lymphoma (OAL), atypical lymphoid hyperplasia (ALH), and reactive lymphoid hyperplasia (RLH) in the Chinese population and its role in clinical diagnosis and pathogenesis. Forty-three samples were collected during patient surgeries. Pathological diagnosis confirmed OAL in 23 cases, ALH in 10 cases, and RLH in 10 cases. Normal peripheral lymph tissues from 12 cases were used as negative controls. Bcl10 gene expression was examined using molecular biological methods, and DNA sequences and mutations were compared with published data. The protein expression of Bcl10 and nuclear factor kappaB (NF-κB) were detected with immunohistological and immunofluorescence colocalization. Bcl10 gene expression was detected in 15 OAL cases. Novel mutations were found in 11 cases. Notably, 1 mutation, which matched a published mutation, was detected in 1 ALH case; 1 novel mutation was found in 1 RLH case; and no Bcl10 gene mutation was found in controls. Most novel mutations were truncation mutations, resulting in a truncated protein product of 99 amino acids (compared to the full-length 233 amino acids; GenBank accession No. EF189176). Results of tests for abnormal Bcl10 gene expression in nuclei or cytoplasm were consistent with changes in NF-κB translocation. This report is the first of newly discovered mutations in the Bcl10 gene in the Chinese population. The distribution of the mutations is consistent with and more sensitive than that of the pathological diagnosis. These mutations can be used to identify the stage and clinical characteristics even when morphological changes are absent.

Surface nanocrystallization of Ti-6Al-4V alloy: microstructural and mechanical characterization.

In this study, microstructural and mechanical properties of Ti-6Al-4V alloy, before and after the SMA treatment (SMAT) as well as the duplex SMAT/Nitriding process at different treatment conditions, were investigated in order to deepen the knowledge of these properties for biomedical devices. For that purpose, tribological (wear resistance, coefficient of friction) and mechanical (Vickers microhardness) tests were performed. To carry out the microstructural and surface topographical characterization of the samples, the scanning electron microscopy (SEM) and the 3D-SEM reconstruction from stereoscopic images have been used. By means of profiles deduced from the 3D images, the surface roughness has been calculated. The obtained results allowed to find an interesting SMAT condition which, followed by nitriding at low temperature, can greatly improve tribological and mechanical properties of Ti-6Al-4V alloy. It was also shown from SEM characterization and the original method of 3D-SEM reconstruction, that SMAT can reduce the machined grooves and consequently the roughness of the samples decreases. Moreover, we demonstrated, for the first time, that instead of usual etching method, the ionic polishing allowed to reveal the grains, the grain boundaries and the twins as well as the surface nanocrystalline layer generated by SMAT. Thus, the thickness of the SMATed layer decreases with the nitriding temperature, whereas the surface grain size increases.

Molecular cloning and mRNA expression profiling of the first specific jasmonate biosynthetic pathway gene allene oxide synthase from Hyoscyamus niger.

In the endeavor to enhance the production of pharmaceutically valuable tropane alkaloids including hyoscyamine and scopolamine in Hyoscyamus niger, methyl jasmonate (MeJA) showed significant stimulation both in tropane biosynthetic pathway enzymes activities and tropane alkaloids yields. Therefore it was speculated that genetic engineering of jasmonate biosynthetic pathway might enhance the endogenous jasmonate concentration, followed by stimulating the production of tropane alkaloids. Herein a full-length cDNA encoding allene oxide synthase (AOS, EC 4.2.1.92), the first committed step enzyme in jasmonate biosynthetic pathway was reported (named HnAOS, GenBank accession: EF532599). HnAOS was a novel member of the cytochrome P450 (CYP74A) subfamily. Real-time quantitative PCR analysis showed that HnAOS mRNA accumulated mainly in stems, and responded significantly to wounding or methyl jasmonate.

Antibiotic-induced alterations of the gut microbiota and microbial fermentation in protein parallel the changes in host nitrogen metabolism of growing pigs.

Gut microbes, especially those in the large intestine, are actively involved in nutrient metabolism; however, their impact on host nitrogen (N) metabolism remains largely unknown. This study was designed to investigate the effects of feeding a cocktail of antibiotics (AGM) (ampicillin, gentamycin and metronidazole) on intestinal microbiota, N utilization efficiency, and amino acid (AA) digestibility in cannulated pigs, with the aim of exploring the impact of gut microbiota on host N metabolism. In total, 16 piglets were surgically fitted with a simple distal ileal T-cannula and a jugular venous catheter. The pigs were fed a basal diet without antibiotics (control; CON) or with antibiotics (antibiotic; ANTI), for 2 weeks. The results showed that feeding AGM did not affect weight gain or digestive enzyme activity. The antibiotics increased the concentration of urea N (P<0.05). However, they reduced N utilization, and the total tract apparent digestibility of isoleucine, methionine, valine, tyrosine and total AA (P<0.05). Furthermore, the antibiotics increased the terminal ileum apparent digestibility of CP, phenylalanine, valine, alanine, tyrosine and total AA (P<0.05). AGM markedly altered the composition of the microbiota in the ileum and feces, with a reduction in populations of Bifidobacterium, Lactobacillus and Ruminococcus, and an increase in the abundance of Escherichia coli (P<0.05). The antibiotics also significantly increased the concentration of cadaverine and ammonia, both in ileal digesta and feces (P<0.05), suggesting a marked impact on N metabolism in the intestine. The analyses indicated that the alteration of gut microbiota was correlated with the apparent digestibility of CP and AA in the intestine. These findings suggest that the AGM-induced alteration of gut microbiota may contribute to the change in intestinal N metabolism, and consequently, N excretion from the body. These results also suggest that antibiotics could have a significant effect on host N metabolism. The present study contributes to our understanding of the effects of antibiotics and provides a rational scientific basis for diet formulation during AGM use.

Rubber seed oil and flaxseed oil supplementation alter digestion, ruminal fermentation and rumen fatty acid profile of dairy cows.

Rubber seed oil (RO) that is rich in polyunsaturated fatty acids (FA) can improve milk production and milk FA profiles of dairy cows; however, the responses of digestion and ruminal fermentation to RO supplementation in vivo are still unknown. This experiment was conducted to investigate the effect of RO and flaxseed oil (FO) supplementation on nutrients digestibility, rumen fermentation parameters and rumen FA profile of dairy cows. Forty-eight mid-lactation Holstein dairy cows were randomly assigned to one of four treatments for 8 weeks, including basal diet (CON) or the basal dietary supplemented with 4% RO, 4% FO or 2% RO plus 2% FO on a DM basis. Compared with CON, dietary oil supplementation improved the total tract apparent digestibility of DM, neutral detergent fibre and ether extracts ( P < 0.05). Oil treatment groups had no effects on ruminal digesta pH value, ammonia N and microbial crude protein ( P > 0.05), whereas oil groups significantly changed the volatile fatty acid (VFA) profile by increasing the proportion of propionate whilst decreasing total VFA concentration, the proportion of acetate and the ratio of acetate to propionate ( P < 0.05). However, there were no differences in VFA proportions between the three oil groups (P > 0.05). In addition, dietary oil supplementation increased the total unsaturated FA proportion in the rumen by enhancing the proportion of trans-11 C18:1 vaccenic acid (VA), cis-9, trans-11 conjugated linoleic acid (CLA) and α-linolenic acid (ALA) ( P < 0.05). These results indicate that dietary supplementation with RO and FO could improve nutrients digestibility, ruminal fermentation and ruminal FA profile by enhancing the VA, cis-9, trans-11 CLA and ALA composition of lactating dairy cows. These findings provide a theoretical basis for the application of RO in livestock production.

[Molecular cloning and expression profile of a jasmonate biosynthetic pathway gene allene oxide cyclase from Hyoscyamus niger].

Hyoscyamus niger L. is a medicinal plant which produces a class of jasmonate-responsive pharmaceutical secondary metabolites named as tropane alkaloids. As a family of signaling phytohormones, jasmonates play significant roles in the biosynthesis of many plant secondary metabolites. In jasmonate biosynthetic pathway of plants, allene oxide cyclase (AOC, [...] EC 5.3.99.6 [...]) catalyzes the most important step. Here we cloned a cDNA from H. niger, named HnAOC (GenBank accession: AY708383), which was 1044 bp long, with a 747 bp open reading frame (ORF) encoding a polypeptide of 248 amino acid residues. Southern blot analysis indicated that it was a multi-copy gene. RT-PCR analysis revealed that the expression of HnAOC was regulated by various stresses and elicitors, with methyl-jasmonate showing the most prominent inducement. The characterization of HnAOC would be helpful for improving the production of valuable secondary metabolites by regulating the biosynthesis ofjasmonates.

[Construction and characterization of normalized cDNA library of maize inbred Mo17 from multiple tissues and developmental stages].

Comprehensive complementary DNA (cDNA) library is a valuable resource for functional genomics. In this study, we set up a normalized cDNA library of Mo17 (MONL) by saturation hybridization with genomic DNA, which contained expressed genes of eight tissues and organs from inbred Mo17 of maize (Zea mays L.). In this library, the insert sizes range from 0.4 kb to 4 kb and the average size is 1.18 kb. 10.830 clones were spotted on nylon membrane to make a cDNA microarray. Randomly picked 300 clones from the cDNA library were sequenced. The cDNA microarry was hybridized with pooled tissue mRNA probes or housekeeping gene cDNA probes. The results showed the normalized cDNA library comprehensively includes tissue-specific genes in which 71% are unique ESTs (expressed sequence tags) based on the 300 sequences analyzed. Using BLAST program to compare the sequences against online nucleotide databases, 88% sequences were found in ZmDB or NCBI, and 12% sequences were not found in existing nucleotide databases. More than 73% sequences are of unknown function. The library could be extensively used in developing DNA markers, sequencing ESTs, mining new genes, identifying positional cloning and candidate gene, and developing microarrays in maize genomics research.

[Isolation and expression profiling of the Pto-like gene SsPto from Solanum surattense].

A novel Pto-like gene (designated as SsPto) is cloned from yellow-fruit nightshade (Solanum surattense). The full-length cDNA of SsPto is 1331 bp long with an open reading frame of 960 bp encoding a polypeptide of 320 amino acid residues. The deduced SsPto protein has a calculated molecular weight of 36.21 kDa with an isoelectric point of 6.18. Multiple sequence alignment shows that SsPto protein shares 71.4% and 71.6% identities to Pto proteins from Lycopersicon pimpinellifolium and L. hirsutum respectively. Genomic Southern blot analysis indicates the presence of a small family of SsPto in the S. surattense genome. SsPto is found to be constitutively expressed in the S. surattense plant with the highest expression in stems. However, under induction by TMV for 6 days, SsPto expresses the highest in roots. Further expression analysis reveals that the signaling components of defense/stress pathways, such as methyl jasmonate (MeJA), salicylic acid (SA), gibberellic acid (GA3) and hydrogen peroxide (H2O2), up-regulate the SsPto transcript levels over the control. Cold treatment, nevertheless, has no significant effect on SsPto expression whereas SsPto expression is down-regulated by dark treatment. Our findings suggest that this novel stress- and pathogen-inducible SsPto from S. surattense may participate not only in the defense/stress responsive pathways, but also in diverse processes of plant's growth and development.