High-Level Extracellular Production of a Trisaccharide-Producing Alginate Lyase AlyC7 in Escherichia coli and Its Agricultural Application.

Alginate oligosaccharides (AOS), products of alginate degradation by endotype alginate lyases, possess favorable biological activities and have broad applications. Although many have been reported, alginate lyases with homogeneous AOS products and secretory production by an engineered host are scarce. Herein, the alginate lyase AlyC7 from Vibrio sp. C42 was characterized as a trisaccharide-producing lyase exhibiting high activity and broad substrate specificity. With PelB as the signal peptide and 500 mM glycine as the additive, the extracellular production of AlyC7 in Escherichia coli reached 1122.8 U/mL after 27 h cultivation in Luria-Bertani medium. The yield of trisaccharides from sodium alginate degradation by the produced AlyC7 reached 758.6 mg/g, with a purity of 85.1%. The prepared AOS at 20 µg/mL increased the root length of lettuce, tomato, wheat, and maize by 27.5%, 25.7%, 9.7%, and 11.1%, respectively. This study establishes a robust foundation for the industrial and agricultural applications of AlyC7.

Cost-effective production of alginate oligosaccharides from Laminaria japonica roots by Pseudoalteromonas agarivorans A3.

BACKGROUND: Alginate oligosaccharides (AOs) are the degradation products of alginate, a natural polysaccharide abundant in brown algae. AOs generated by enzymatic hydrolysis have diverse bioactivities and show broad application potentials. AOs production via enzymolysis is now generally with sodium alginate as the raw material, which is chemically extracted from brown algae. In contrast, AOs production by direct degradation of brown algae is more advantageous on account of its cost reduction and is more eco-friendly. However, there have been only a few attempts reported in AOs production from direct degradation of brown algae. RESULTS: In this study, an efficient Laminaria japonica-decomposing strain Pseudoalteromonas agarivorans A3 was screened. Based on the secretome and mass spectrum analyses, strain A3 showed the potential as a cell factory for AOs production by secreting alginate lyases to directly degrade L. japonica. By using the L. japonica roots, which are normally discarded in the food industry, as the raw material for both fermentation and enzymatic hydrolysis, AOs were produced by the fermentation broth supernatant of strain A3 after optimization of the alginate lyase production and hydrolysis parameters. The generated AOs mainly ranged from dimers to tetramers, among which trimers and tetramers were predominant. The degradation efficiency of the roots reached 54.58%, the AOs production was 33.11%, and the AOs purity was 85.03%. CONCLUSION: An efficient, cost-effective and green process for AOs production directly from the underutilized L. japonica roots by using strain A3 was set up, which differed from the reported processes in terms of the substrate and strain used for fermentation and the AOs composition. This study provides a promising platform for scalable production of AOs, which may have application potentials in industry and agriculture.

Study of pathogenic genes in a pedigree with familial dilated cardiomyopathy.

BACKGROUND: Dilated cardiomyopathy (DCM) is a genetically heterogeneous cardiac disorder characterized by left ventricular dilation and contractile dysfunction. The substantial genetic heterogeneity evident in patients with DCM contributes to variable disease severity and complicates overall prognosis, which can be very poor. AIM: To identify pathogenic genes in DCM through pedigree analysis. METHODS: Our research team identified a patient with DCM in the clinic. Through investigation, we found that the family of this patient has a typical DCM pedigree. High-throughput sequencing technology, next-generation sequencing, was used to sequence the whole exomes of seven samples in the pedigree. RESULTS: A novel and potentially pathogenic gene mutation-ANK2p.F3067L-was discovered. The mutation was completely consistent with the clinical information for this DCM pedigree. Sanger sequencing was used to further verify the locus of the mutation in pedigree samples. These results were consistent with those of high-throughput sequencing. CONCLUSIONS: ANK2p.F3067L is considered a novel and potentially pathogenic gene mutation in DCM.

Iron-catalyzed regioselective synthesis of (E)-vinyl sulfones mediated by unprotected hydroxylamines.

An Fe-catalyzed unprotected hydroxylamine mediated Heck-type coupling between sulfinic acids and alkenes for the regioselective synthesis of (E)-vinyl sulfones has been developed. Mechanism studies indicated for the first time that a radical process may be involved and that hydroxylamines play multiple roles, including those of a mild oxidant and an in situ base. It was found for the first time that this transformation not only realizes C-S bond construction promoted by unprotected hydroxylamines, but also provides a practical and complementary method for the preparation of structurally important (E)-vinyl sulfones.

[Chemical Forms and Spatial Distribution of Phosphorus in the Sediment of Sihe River].

Based on the study of the content, forms, and spatial distribution of phosphorus (P) in the surface and columnar sediments of the Sihe River, the relationships between the total phosphorus (TP) and various forms of P and the basic physical and chemical properties of sediment and their ecological significance were deeply discussed. The forms of P in the sediment were defined including soluble and loosely bound P (S/L-P), Al-bound P (Al-P), Fe-bound P (Fe-P), reductant soluble P (RS-P), Ca-bound P (Ca-P), and residual P (RES-P) using the method of selectively sequential extraction. The results indicated that ω (TP) was 421.84-1188.65 mg·kg-1 in the surface sediment. Among the six forms of P, the content of Ca-P was the highest, accounting for more than 40% of TP, followed by the content of Fe-P, accounting for more than 20% of TP. The content of S/L-P was the lowest, which only accounted for approximately 0.4% of TP. The contents of TP and various forms of P in the surface sediments of the downstream area of the Sihe River were higher than those of the upstream area of the river, which was induced by the discharge of industrial wastewater and domestic sewage in the urban areas nearby the downstream portion of the Sihe River. The contents of TP in the upper samples of the two sediment profiles were obviously higher than those in the bottom samples, indicating that the P pollution in the water environment of the Sihe River has been intensifying in recent years. Among all forms of P in the sediment profiles, Ca-P accounted for the largest proportion, followed by Res-P. The correlation analysis results showed that significant correlations were observed between Fe and Fe-P, Al and Al-P, Ca and Ca-P, and TOC and RS-P in the surface sediments; the same correlations had not been found in the sediment profiles. The calculated results of Fe and TP molar ratio indicated that the sediments of the Sihe River could further accumulate P. The percentage of bioavailable P (BAP) in sediment was 25%-50%.

[GAO Shu-zhong's clinical experience in "seeking yin from yang needling method" based on "qi street" and "four seas" theories].

The paper introduces professor GAO Shu-zhong's understanding on "seeking yin from yang needling method" and its clinical application on the basis of "qi street" and "four seas" theories. Through professor GAO's clinical practice for years, he integrates and extendes the theories of "seeking yin from yang", "qi street" and "four seas" in Huangdi Neijing (The Yellow Emperor's Inner Classic). In this specific acupuncture method, in reference with the theories of "qi street" and "four seas", acupuncture is exerted on yang part of body, e.g. the back and lumber region to treat the diseases of yin parts, e.g. the chest and abdomen, which is differentiated as yin-yang imbalance in pathogenesis. In order to fully explain the clinical curative effect of "seeking yin from yang needling method", the common diseases in clinic, e.g. the disorders of heart, spleen and stomach systems, as well as the gynecology are taken as examples in the paper.

Comparative analysis unveils the cadmium-induced reproductive toxicity on the testes of Pardosa pseudoannulata.

Cadmium (Cd) pollution is one of the most serious heavy metal pollutions in the world, which has been demonstrated to cause different toxicities to living organisms. Cd has been widely suggested to cause reproductive toxicity to vertebrates, yet its reproductive toxicity to invertebrates is not comprehensive. In this study, the wolf spider Pardosa pseudoannulata was used as a bioindicator to evaluate the male reproductive toxicity of invertebrates under Cd stress. Cd stress had no effect on the color, size and length of testis. However, Cd significantly increased the contents of catalase, glutathione peroxidase and malondialdehyde, the antioxidants in the testis of P. pseudoannulata. Then we analyzed the transcriptome of testis exposed to Cd, and identified a total of 4739 differentially expressed genes (DEGs) compared to control, with 2368 up-regulated and 2371 down-regulated. The enrichment analysis showed that Cd stress could affect spermatogenesis, sperm motility, post-embryonic development, oxidative phosphorylation and metabolism and synthesis of male reproductive components. At the same time, the protein-protein interaction network was constructed with the generated DEGs. Combined with the enrichment analysis of key modules, it revealed that Cd stress could further affect the metabolic process in testis. In general, the analysis of testicular damage and transcriptome under Cd stress can provide a novel insight into the reproductive toxicity of Cd on rice filed arthropods and offer a reference for the protection of rice filed spiders under Cd pollution.

Icaritin-elevated circ_0000190 suppresses the malignant progression of multiple myeloma by targeting miR-301a.

Icaritin has potential anticancer effects on various cancers, including multiple myeloma (MM). Recent studies claim that Icaritin can regulate the expression of noncoding RNAs (ncRNAs) in cancer development. This study aimed to investigate the role of circular RNA_0000190 (circ_0000190) and functional mechanism in Icaritin-treated MM. The expression of circ_0000190 and miR-301a was detected by quantitative real-time polymerase chain reaction. Cell cycle, apoptosis, migration, and invasion were investigated using flow cytometry assay, and transwell assay, respectively. The expression of BAX, BCL2, MMP2, and CCND1 was detected by western blot. The predicted target relationship between circ_0000190 and miR-301a was validated by dual-luciferase reporter assay and RNA immunoprecipitation assay. The activation of JAK1/STAT3 pathway was examined using western blot. Circ_0000190 was strikingly downregulated in MM specimens and cell lines, and Icaritin promoted the expression of circ_0000190. In function, circ_0000190 overexpression promoted MM cell cycle arrest and apoptosis but restrained the ability of migration and invasion. Icaritin blocked the development of MM by increasing circ_0000190 expression. MiR-301a was identified as a target of circ_0000190, and miR-301a reintroduction largely abolished the effects of circ_0000190 overexpression. The activation of JAK1/STAT3 pathway was promoted by miR-301a restoration. Icaritin played anticancer effects in MM partly by enhancing the expression of circ_0000190 and regulating the circ_0000190/miR-301a pathway. This study enhanced the understanding of the mechanism of Icaritin associated with circRNAs in MM.

Effectiveness of Solifenacin for Managing of Bladder Spasms in Patients With Urethroplasty.

To evaluate the efficacy and safety of solifenacin in the treatment of bladder spasms after urethroplasty. Patients underwent urethroplasty were randomly assigned to the study group ( n = 165) and the control group ( n = 150). Patients in the study group were treated with solifenacin for 7 days. Patients in the control group were placebo. Each group was further divided into four subgroups: paracentetic suprapubic cystostomy subgroup, traditional suprapubic cystostomy subgroup, former suprapubic cystostomy subgroup, and urethral catheter subgroup. A visual analogue scale (VAS) was used to measure the severity of bladder spasms. The mean duration of spasms, the frequency of spasms, and the incidences of urine extravasation and radiating pain were recorded each day. There were no significant differences in the VAS scores and mean duration of bladder spasms between the study and control groups . However, there was a significantly lower VAS score in the patients taking solifenacin in the paracentetic suprapubic cystostomy subgroup ( p < .05). A similar tendency was noted in the mean duration of bladder spasms in this subgroup. In a comparison of the daily and nightly frequency of spasms within the four subgroups, a significant improvement was noted in the control group within 5 days. A similar difference was not noted within 6 days in the study group. The short-term therapy with solifenacin is an effective and safe method for decreasing the frequency of bladder spasms after urethroplasty. Patients undergoing paracentetic suprapubic cystostomy might be the only subset to benefit from this treatment.

Salidroside-regulated lipid metabolism with down-regulation of miR-370 in type 2 diabetic mice.

Salidroside is known for its pharmacological properties and in particular its antioxidation effects. In recent years, it has been recognized that salidroside plays an important role in treating diabetes. Accumulated evidence suggests that microRNAs may be involved in diabetic lipid disorders. We investigated how salidroside regulates lipid metabolism through miR-370 in vivo and in vitro. After 4 weeks of a high-fat diet, and intraperitoneal injection of streptozotocin (100mg/kg), type 2 diabetes was induced in male C56BL/6J mice. After 4 weeks, mice with fasting blood glucose levels above 7.8mmol/l were divided into five groups: those with diabetes mellitus, and those treated with 40mg/kg, 80mg/kg, and 160mg/kg salidroside, and metformin (480mg/kg), for a further 4 weeks. The hypoglycemic effects of salidroside were consistently demonstrated when measuring fasting blood glucose levels, observing insulin-sensitizing effects, and testing oral glucose tolerance. In addition to this, the expressions of miR-370, and related lipid protein expression in primary hepatocytes, were examined in primary type 2 diabetic mice. The present study has shown that the expression levels of miR-370, SREBP-1 and FAS-1 were significantly elevated in the liver of type 2 diabetic mice. In contrast, the elevated expression levels were reversed by salidroside. The addition of salidroside attenuated the effect of miR-370, and reduced the expression of these lipid metabolism proteins in primary hepatocytes. These findings demonstrate that salidroside can directly decrease the expression of miR-370 in type 2 diabetic mice, and particularly in primary hepatocytes, affecting lipid metabolism in the liver.

Long-term Outcome of Ileal Ureteric Replacement With an Iliopsoas Muscle Tunnel Antirefluxing Technique for the Treatment of Long-segment Ureteric Strictures.

OBJECTIVE: To investigate the long-term outcome of ileal ureteric replacement using a proximal antirefluxing technique for the treatment of long-segment ureteric strictures. PATIENTS AND METHODS: Between January 1997 and December 2013, 41 patients with a long ureteral stricture or defect and 3 patients with unilateral mid-lower ureteral cancer (20 bilateral and 24 unilateral, 28 males and 16 females) were treated by ureteral substitution using a proximal antirefluxing technique. The distal part of the upper ureter (4 cm) was fixed between the psoas muscle and the ileal segment (the iliopsoas tunnel technique). The distal ileum was connected to the urinary bladder with an end-to-side anastomosis. A successful outcome was defined as the absence of major complications, worsening baseline renal function, metabolic derangements, or obstruction. RESULTS: One patient with unilateral mid-lower ureteral cancer died 3 years postoperatively because of metastasis, and the remaining 43 patients were followed for 12-180 months (mean 69 months). Intravenous urography showed that the hydronephrosis improved significantly or disappeared after 6-12 months in 34 patients, with improvement in 9 patients. Cystography showed no evidence of ileoureteral reflux. Seven patients needed long-term oral alkalization to prevent hyperchloremic acidosis. CONCLUSION: In our experience, outcomes following subtotal ureteric replacement are encouraging. The ileal ureter replacement by the proximal antirefluxing technique appears to be a reliable procedure for treating long-segment ureteral stricture and preservation of renal function.

Two Micrometer Continuous-Wave Thulium Laser Treating Primary Non-Muscle-Invasive Bladder Cancer: Is It Feasible? A Randomized Prospective Study.

BACKGROUND AND OBJECTIVE: Until now, various laser types have been used in the treatment of bladder cancer. The purpose of this study is to evaluate the feasibility and effectiveness of 2 µm continuous-wave (CW) thulium laser in treating primary non-muscle-invasive bladder cancer (NMIBC). METHODS: A comparative (randomized prospective) interventional study was conducted in our center. A total of 400 patients with Ta or T1 bladder cancer were enrolled and randomized to transurethral resection of bladder tumor (TURBT) or laser vaporesection (LVRBT). Patients in the TURBT group received standard transurethral resection with a bipolar electrocautery system, while those in the LVRBT group received transurethral vaporesction with a 2 µm CW thulium laser. All patients received intravesical irrigation with sterile water for at least 12 h immediately after the procedure. The end-points of this study were tumor recurrence and/or progression, or the completion of 36 months of follow-up (10 cystoscopies) with no recurrence. RESULTS: A total of 292 patients were included in our analysis, of whom 143 received TURBT and 149 received LVRBT. During 36 months of follow-up, 129 cases (44.2%) had recurrence: 61 (42.7%) in the TURBT group and 68 (45.6%) in the LVRBT group (p = 0.608). Tumor progression was found in 19 patients (6.5%): 11 in the TURBT (7.7%) and 8 (5.4%) in the LVRBT group (p = 0.421). Median tumor-free survival was 25.2 months (25.46 in the TURBT group vs. 24.9 in the LVRBT group, p = 0.729). Overall recurrence rate was 14.7%, 42.1%, and 62.5% in the low, intermediate low, and intermediate high risk subgroups, respectively. Operation time was significantly lower in the TURBT than in the LVRBT group (28.43 vs. 31.5 min, p = 0.044). CONCLUSIONS: Use of a 2 µm CW thulium laser is feasible and effective in treating primary NMIBC. With a cystoscopic biopsy of the bladder muscle, 2 µm CW thulium laser could retrieve sufficient diagnostic and prognostic specimens.

Pedicled rectus abdominis muscle and fascia flap sling the bulbar urethra for treatment for male-acquired urinary incontinence: report of ten cases.

PURPOSE: Male urinary incontinence is relatively common complication of radical prostatectomy and of posterior urethroplasty following traumatic pelvic fracture. Here, we investigate the use of pedicled rectus abdominis muscle and fascia flap sling of the bulbar urethra for treatment for male-acquired urinary incontinence. MATERIALS AND METHODS: Ten patients with acquired urinary incontinence were included in the study. Urinary incontinence was secondary to TURP in three patients and was secondary to posterior urethroplasty performed following traumatic pelvic fracture in seven patients. Pedicled rectus abdominalis muscle and fascial flaps, approximately 2.5 cm wide and 15 cm long, were isolated. The flaps were inserted into a perineal incision through a subcutaneous tunnel. The free end of the flap was sectioned to form two muscle strips, each 3 cm in length, and inserted into the space between bulbar urethra and corpus cavernosa. After adequate sling tension had been achieved, the two strips of muscle were anastomosed around the bulbar urethra using a 2-zero polyglactin suture. RESULTS: The patients were followed up for between 12 and 82 months (mean 42.8 months). Complete continence was achieved with good voiding in seven of the 10 patients. In other three patients achieved good voiding following catheter removal, but incontinence was only moderately improved. CONCLUSIONS: A pedicled rectus muscle fascial sling of the bulbar urethra is an effective and safe treatment for male patients with mild to moderate acquired urinary incontinence, but it may not be suitable for severe incontinence or for patients with weak rectus abdominalis muscles.

[Systemic oxidative stress induced by intratracheal instilling with PM10 in rats].

OBJECTIVE: To observe the oxidative stress of serum, lung and heart tissues in rats caused by PM10. METHODS: In the study,24 male Wistar rats weighed 320-360 g were randomly divided into four groups, including one saline control group and three PM10 exposed groups. PM10 was administered to the exposed groups by intratracheal instillation at the doses of 3.75, 7.5, 15 mg/kg, respectively. The rats were sacrificed 24 hours after exposure. The levels of MDA and activities of SOD in the serum, lung and heart tissues were measured. In addition, the contents of protein carbonyl in lung and heart tissues were determined. RESULTS: The activities of SOD in the serum, lung and heart tissues decreased markedly in the 7.5 and 15 mg/kg groups. The levels of MDA in the lung tissues significantly increased in the 7.5 and 15 mg/kg groups, while those in the serum and heart tissues significantly increased only in the 15 mg/kg group. The contents of protein carbonyl in the lung tissues markedly increased in the 15 mg/kg group, but no significant change was found in the heart tissues. CONCLUSION: The study showed that PM10 can cause oxidative stress effects in rats. The lipids of cell membrane are more susceptible to oxidative stress than the proteins.

Reinforcing the continent mechanism of continent cutaneous diversions by wrapped rectus abdominis muscle flap: a preliminary experimental study.

BACKGROUND: Continent cutaneous diversion (CCD) has been widely used in almost any lower urinary reconstruction. We have been continually trying to modify this procedure because of the high complications rate, especially as they relate to the efferent tube. In this study, we reported a modified procedure with a tapered ileum wrapped by the rectus abdominalis flap (RAMF) and assessed the feasibility of this new technique to achieve urinary continence. METHODS: A procedure in which two ileal segments were tapered and connected to a U-shaped reservoir was performed in ten dogs. A RAMF with its blood supply was wrapped around one of the tapered ileum. In control groups, the tapered ileum was brought to the abdominal skin. Urodynamic studies were conducted In the 1st, 3rd and 6th months post-operatively. The data of maximum inner pressure (MIP) and functional pressure length (FPL) in every group at each phase were recorded. Retrograde radiograms of the efferent limbs were performed before sacrifice. RESULTS: MIP in the study group was significantly higher than that in the control group at each phase (P < 0.05). However, no significant differences in MIP or FPL were found in the study group between an empty and full reservoir. In the control group, MIP increased (P < 0.05) and FPL decreased significantly (P < 0.05) compared with an empty and full reservoir. Retrograde radiograms confirmed that efferent limbs were positioned straigh beneath the abdominal wall. Histological examination of the study group demonstrated a layer of striated muscle around the outside surface of the ileum. CONCLUSION: The continent mechanism of tapered ileum can be enhanced by extra support from wrapped RAMF.

[Reconstructive surgery for male urethral stricture affects erectile function and sexual life quality].

OBJECTIVE: To evaluate the influence of reconstructive surgery for male urethral stricture on erectile function and sexual life quality. METHODS: We analyzed retrospectively the clinical data of 326 male patients who underwent urethroplasty for urethral stricture in our department and evaluated their erectile function and sexual life quality. RESULTS: A total of 172 groups of valid data were collected, with the mean follow-up of 28.5 months. The mean scores on IIEF-5 (P=0.002) and sexual life quality (P=0.026) were statistically significantly reduced after surgery. Erectile dysfunction was found in 88 (51.2%) of the patients after urethroplasty, as compared with 56 (32.6%) preoperatively. CONCLUSION: The location of urethral stricture, surgical method and urethral stricture recurrence may affect the erectile function and sexual life quality of the patient, but both can be gradually improved with the time after urethroplasty.

Ileal ureteric replacement with an ileo-psoas muscle tunnel antirefluxing technique for the treatment of long segment ureteric strictures.

OBJECTIVES: To develop and report our initial experience with a novel antirefluxing technique for segmental ileal ureteric replacement for the treatment of long ureteric strictures. PATIENTS AND METHODS: Between January 2000 and January 2007, 12 cases of ureteric strictures (nine bilateral and three unilateral) were treated using a novel surgical technique in which the ureter was replaced with a segment of ileum using an end-to-side anastomosis. An antireflux valve was constructed by fixing the distal part of upper ureter (4 cm) between the psoas muscle and ileal segment (the ileo-psoas tunnel technique). The distal ileum was connected to the urinary bladder with an end-to-side anastomosis. RESULTS: The 12 cases were followed-up for a mean (range) of 39.42 (12-64) months. There were no cases of pyelonephritis or signs of renal failure after surgery. There was dramatic improvement in hydronephrosis, as assessed by intravenous urography, in the 4-9 months after surgery. Cystography showed no evidence of ileo-ureteric reflux. Mild hyperchloraemic acidosis was detected in two patients and was successfully treated with oral alkalization. CONCLUSIONS: In our initial experience, ileal ureteric replacement combined with the ileo-psoas tunnel antirefluxing technique is a highly effective procedure for the treatment of ureteric strictures.

[Etiopathogenesis and management of seminal vesicle enlargement].

OBJECTIVE: To study the etiopathogenesis and management of seminal vesicle enlargement. METHODS: Forty-six cases of seminal vesicle were selected, of which 36 were vesiculitis, treated by antibiotics through a catheter indwelt in the seminal vesicle for 1 week, 3 were cystic vesicular seminalis, given anti-inflammatory treatment by catheter administration and hydatid fluid inhalation, 1 was calculus at the orifice of the ejaculatory duct, taken out by transurethral resection of the verumontanum, 1 was polypous at the orifice of the verumontanum, removed by transurethral resection, 2 were posterior urethritis, treated by fulguration, and 3 were prostatic carcinoma, treated by radical prostatectomy and spermatocystotomy. RESULTS: Hemospermia disappeared in 32 cases of vesiculitis during the 6-24 months follow-up, 4 cases experienced recurrent hemospermia 3 months after the treatment. No recurrence was observed in any other case of cystic vesicular seminalis, calculus, polypous, posterior urethritis and prostatic carcinoma. CONCLUSION: Seminal vesicle enlargement has intricate etiopathogenesis, but can be treated with satisfactory results if managed properly.