Astragaloside IV Alleviates Podocyte Injury in Diabetic Nephropathy through Regulating IRE-1α/NF-κ B/NLRP3 Pathway.

OBJECTIVE: To investigate the effects of astragaloside IV (AS-IV) on podocyte injury of diabetic nephropathy (DN) and reveal its potential mechanism. METHODS: In in vitro experiment, podocytes were divided into 4 groups, normal, high glucose (HG), inositol-requiring enzyme 1 (IRE-1) α activator (HG+thapsigargin 1 µmol/L), and IRE-1α inhibitor (HG+STF-083010, 20 µmol/L) groups. Additionally, podocytes were divided into 4 groups, including normal, HG, AS-IV (HG+AS-IV 20 µmol/L), and IRE-1α inhibitor (HG+STF-083010, 20 µmol/L) groups, respectively. After 24 h treatment, the morphology of podocytes and endoplasmic reticulum (ER) was observed by electron microscopy. The expressions of glucose-regulated protein 78 (GRP78) and IRE-1α were detected by cellular immunofluorescence. In in vivo experiment, DN rat model was established via a consecutive 3-day intraperitoneal streptozotocin (STZ) injections. A total of 40 rats were assigned into the normal, DN, AS-IV [AS-IV 40 mg/(kg·d)], and IRE-1α inhibitor [STF-083010, 10 mg/(kg·d)] groups (n=10), respectively. The general condition, 24-h urine volume, random blood glucose, urinary protein excretion rate (UAER), urea nitrogen (BUN), and serum creatinine (SCr) levels of rats were measured after 8 weeks of intervention. Pathological changes in the renal tissue were observed by hematoxylin and eosin (HE) staining. Quantitative reverse transcription-polymerase chain reaction (RT-PCR) and Western blot were used to detect the expressions of GRP78, IRE-1α, nuclear factor kappa Bp65 (NF-κBp65), interleukin (IL)-1ß, NLR family pyrin domain containing 3 (NLRP3), caspase-1, gasdermin D-N (GSDMD-N), and nephrin at the mRNA and protein levels in vivo and in vitro, respectively. RESULTS: Cytoplasmic vacuolation and ER swelling were observed in the HG and IRE-1α activator groups. Podocyte morphology and ER expansion were improved in AS-IV and IRE-1α inhibitor groups compared with HG group. Cellular immunofluorescence showed that compared with the normal group, the fluorescence intensity of GRP78 and IRE-1α in the HG and IRE-1α activator groups were significantly increased whereas decreased in AS-IV and IRE-1α inhibitor groups (P<0.05). Compared with the normal group, the mRNA and protein expressions of GRP78, IRE-1α, NF-κ Bp65, IL-1ß, NLRP3, caspase-1 and GSDMD-N in the HG group was increased (P<0.05). Compared with HG group, the expression of above indices was decreased in the AS-IV and IRE-1α inhibitor groups, and the expression in the IRE-1α activator group was increased (P<0.05). The expression of nephrin was decreased in the HG group, and increased in AS-IV and IRE-1α inhibitor groups (P<0.05). The in vivo experiment results revealed that compared to the normal group, the levels of blood glucose, triglyceride, total cholesterol, BUN, blood creatinine and urinary protein in the DN group were higher (P<0.05). Compared with DN group, the above indices in AS-IV and IRE-1α inhibitor groups were decreased (P<0.05). HE staining revealed glomerular hypertrophy, mesangial widening and mesangial cell proliferation in the renal tissue of the DN group. Compared with the DN group, the above pathological changes in renal tissue of AS-IV and IRE-1α inhibitor groups were alleviated. Quantitative RT-PCR and Western blot results of GRP78, IRE-1α, NF-κ Bp65, IL-1ß, NLRP3, caspase-1 and GSDMD-N were consistent with immunofluorescence analysis. CONCLUSION: AS-IV could reduce ERS and inflammation, improve podocyte pyroptosis, thus exerting a podocyte-protective effect in DN, through regulating IRE-1α/NF-κ B/NLRP3 signaling pathway.

[Effects of astragaloside IV on delaying kidney aging and its mechanisms].

OBJECTIVE: To investigate the mechanisms of Astragaloside Ⅳ on inhibiting apoptosis and delaying kidney aging in rats by regulating SIRT1/p53 signaling pathway. METHODS: The aging model was established by subcutaneous injection of D-galactose 200 mg/(kg·d). SPF-grade healthy male SD rats were randomly divided into 4 groups: normal control group (intragastric infusion of 5 ml/(kg·d) normal saline), aging model group (intragastric infusion of 5 ml/(kg·d) normal saline), Astragaloside IV group (intragastric infusion of 40 mg/(kg·d) Astragaloside IV),and SRT1720 group( intragastric infusion of 20 mg/(kg·d) SRT1720), with 10 rats in each group. After 8 weeks, the serum samples of rats were collected to detect the levels of renal function (creatinine and urea nitrogen) and senescent associated secretory phenotype (TGF-ß and IL-6) by ELISA. The renal tissues of rats were obtained for HE and Masson staining. The protein and mRNA expressions of SIRT1, p53, Bcl-2, Bax, p21 and pRb were detected by Western blot and RT-PCR. RESULTS: Serum creatinine and urea nitrogen levels in the aging model group were higher than those in the normal group, but there was no significant difference in each group (P＞0.05). The serum levels of TGF-ß and IL-6 in the aging model group were higher than those in the normal group (P＜0.05), and which in the Astragaloside IV group and SRT1720 group were lower than those in the model group (P＜0.05). There was no significant differences between Astragaloside IV group and SRT1720 group (P＞0.05). The results of pathological staining of renal tissues showed that, compared with the normal group, the renal tubules dilated, local atrophy, infiltration of inflammatory cells and proliferation of collagen fibers were observed in the aging model group. Compared with the aging model group, the pathological changes were alleviated in Astragaloside IV group and SRT1720 group. The results of Western blot and RT-PCR showed that, compared with the normal group, the protein and mRNA expressions of SIRT1 and pRb in the renal tissue of the aging group were decreased, the protein expression of Bcl-2 was decreased(P＜0.05), and the protein and mRNA expressions of p53 and p21 were increased, the protein expression of Bax was increased(P＜0.05). Compared with the aging group, Astragaloside IV and SRT1720 improved the above-mentioned indexes (P＜0.05). CONCLUSION: Astragaloside IV can delay kidney aging by regulating the SIRT1/p53 signaling pathway.

A comparative analysis of immunogenicity and safety of an enterovirus 71 vaccine between children aged 3-5 years and infants aged 6-35 months.

BACKGROUND: The Sinovac enterovirus 71 (EV71) vaccine has shown good safety, immunogenicity, and efficacy in infants aged 6-35 months, whom are considered as the priority of the target population. However, 3-5 years old children accounted for approximately 30% of HFMD cases and are also worth our attention. METHODS: A randomized, double-blind, placebo-controlled, batch-to-batch consistency clinical trial enrolling 1400 participants aged 6-59 months was performed. We pooled the participants receiving three batches of EV71 vaccine together and then stratified them into the 6-35 months and 3-5 years. The non-inferiority analysis of the geometric mean titer (GMT) of EV71 neutralizing antibody post-vaccination was the primary endpoint. RESULTS: In the vaccine group, the GMT of 242 children aged 3-5 years was 132.72 (95% CI, 110.3-159.6), which was non-inferior to that generated in 717 infants aged 6-35 months. Following the vaccination, the incidence of adverse reactions was less frequent in children aged 3-5 years (47.0%) than that found in infants aged 6-35 months (60.1%) (p = 0.0026). CONCLUSIONS: Our study indicated that the EV71 vaccine was also safe in children aged 3-5 years, with non-inferior immunogenicity to that in infants aged 6-35 months.

Glucose deprivation induces chemoresistance in colorectal cancer cells by increasing ATF4 expression.

AIM: To investigate the role of activating transcription factor 4 (ATF4) in glucose deprivation (GD) induced colorectal cancer (CRC) drug resistance and the mechanism involved. METHODS: Chemosensitivity and apoptosis were measured under the GD condition. Inhibition of ATF4 using short hairpin RNA in CRC cells under the GD condition and in ATF4-overexpressing CRC cells was performed to identify the role of ATF4 in the GD induced chemoresistance. Quantitative real-time RT-PCR and Western blot were used to detect the mRNA and protein expression of drug resistance gene 1 (MDR1), respectively. RESULTS: GD protected CRC cells from drug-induced apoptosis (oxaliplatin and 5-fluorouracil) and induced the expression of ATF4, a key gene of the unfolded protein response. Depletion of ATF4 in CRC cells under the GD condition can induce apoptosis and drug re-sensitization. Similarly, inhibition of ATF4 in the ATF4-overexpressing CRC cells reintroduced therapeutic sensitivity and apoptosis. In addition, increased MDR1 expression was observed in GD-treated CRC cells. CONCLUSION: These data indicate that GD promotes chemoresistance in CRC cells through up-regulating ATF4 expression.

Immunogenicity, safety, and lot consistency of a novel inactivated enterovirus 71 vaccine in Chinese children aged 6 to 59 months.

The determination of lot-to-lot consistency in the manufacturing process is a mandatory step in the clinical development of the novel enterovirus 71 (EV71) vaccine. A phase III, randomized, placebo-controlled, double-blind trial assessed the lot consistency, immunogenicity, and safety of the EV71 vaccine in children aged 6 to 59 months. Healthy children (n = 1,400) received one of three lots of the EV71 vaccine containing 400 U of EV71 antigen or a placebo at days 0 and 28. Blood samples were collected before dose 1 and at 28 days after dose 2 (day 56) for an anti-EV71 neutralizing antibody (NTAb) assay. The geometric mean titer (GMT) and the seropositivity rates (with titers of ≥1:8) were compared at day 56. After each dose, the solicited injection site and general adverse events (AEs) were recorded for 7 days, and unsolicited AEs were recorded for 28 days. At day 56, the seropositivity rates ranged from 99.7% to 100% for the vaccine groups. The NTAb GMTs for the vaccine were 140.3 (95% confidence interval [CI], 117.8 to 167.1), 141.5 (95% CI, 118.0 to 169.6), and 146.6 (95% CI, 122.5 to 175.3). The two-sided 95% CI of the log difference in GMTs between the pairs of lots were between -0.176 and 0.176, therefore meeting the predefined equivalence criteria. The percentages of subjects reporting any injection site AEs, general AEs, or serious AEs were similar across the four vaccination groups. In conclusion, the demonstration of consistency between the manufacturing lots confirms for the purposes of clinical development the reliability of the EV71 vaccine production process. (This study has been registered at ClinicalTrials.gov under registration no. NCT01636245.).

Transnasal esophagogastroduodenoscopy for evaluation of upper gastrointestinal non-neoplastic disorders in patients with fresh hypopharyngeal cancer.

OBJECTIVES/HYPOTHESIS: Upper gastrointestinal (UGI) tract non-neoplastic disorders are detrimental to cancer treatment. This study is to evaluate the feasibility of transnasal esophagogastroduodenoscopy in the diagnosis of UGI disorders at the same time as the diagnosis of hypopharyngeal cancer and to provide the prevalence of UGI disorders in patients with fresh hypopharyngeal cancer. STUDY DESIGN: A prospective cases series study. METHODS: Patients with newly diagnosed hypopharyngeal cancer between 2007 and 2010 were enrolled. An endoscope (GIF-XP260N; Olympus Optical, Tokyo, Japan) without conscious sedation was transnasally used to evaluate from the pharynx to duodenum. RESULTS: One hundred two patients were evaluated. Sixty-five patients had at least one UGI disorder: gastric/duodenal ulcers in 32; active Helicobacter pylori infection in 36; and erosive esophagitis in 20 patients, including Los Angeles classification grade A in eight patients, grade B in 10, grade C in one, and grade D in one. Of the abovementioned disorders, 45 patients had only one, 17 patients had two, and three patients had all of the above disorders. Neither age, sex, location, nor T classification of the hypopharyngeal tumor was associated with the presence of UGI disorders. CONCLUSIONS: Transnasal esophagogastroduodenoscopy can be a single procedure to diagnose hypopharyngeal cancer and UGI disorders at one session. Approximately ⅔ of the patients with fresh hypopharyngeal cancer had either erosive esophagitis, active H. pylori infection, or gastric/duodenal ulcers.

Common genetic mutations in the start codon of the SDH subunit D gene among Chinese families with familial head and neck paragangliomas.

Head and neck paragangliomas (HNPGLs) are rare, and frequently associated with germline mutations of the succinate dehydrogenase (SDH) genes, especially for familial cases. The purpose of the study is to explore SDH mutations in Chinese families with familial HNPGLs in Taiwan. Four unrelated families with familial HNPGLs were screened for germline mutations in the SDHB, SDHC and SDHD genes by direct sequencing. One hundred healthy subjects without a diagnosis or family history of HNPGLs were screened as normal controls. Immunohistochemistry with SDHB antibody was performed for a carotid body tumor. Two allele variants were identified, including p.Met1Val (c.1A>G) in the SDHD gene in one family and p.Met1Ile (c.3G>C) in the SDHD gene in the other three families. Both variants are considered pathogenic because of the absence of these variants in 100 normal controls, 100% evolutionary conservation of the p.Met1 residue, co-segregation of the variants with the phenotype of HNPGL in pedigrees, and predicted abolishment of the translation start site. The tumor cells obtained from one proband harboring c.3G>C mis-sense mutation were weak diffuse staining in the cytoplasm of tumors cells. This study demonstrates that two mis-sense mutations at the start codon of the SDHD gene, including p.Met1Val (c.1A>G) and p.Met1Ile (c.3G>C), might be mutation hotspots in Chinese patients with familial HNPGLs.

Salvage treatment for isolated regional failure of nasopharyngeal carcinoma after primary radiotherapy.

BACKGROUND: Isolated regional failure of nasopharyngeal carcinoma (NPC) after primary treatment is traditionally treated with radical neck dissection (RND). The roles of modified radical neck dissection (MRND) and postoperative radiotherapy after salvage surgery currently remain unclear. METHODS: Medical records from our facility of all NPC patients with isolated regional failure after complete primary radiotherapy and receipt of radical surgery as a part of salvage treatment between January 1985 and December 2004 were retrospectively reviewed. RESULTS: Forty-five patients were enrolled onto the study. On univariate analyses, the 5-year regional-free, disease-free, and overall survival rates were 67.7%, 47.8%, and 65.7% for patients who underwent salvage surgery alone and were 66.0%, 34.7%, and 61.3% for patients who received salvage surgery plus postoperative radiotherapy (P = 0.74, P = 0.39 and P = 0.7, respectively). The 5-year regional-free, disease-free, and overall survival rates were 87.4%, 53.5%, and 87.1% for patients undergoing RND and were 54.3%, 34.2%, and 50.5% for patients undergoing MRND (P = 0.01, P = 0.02 and P = 0.05, respectively). On multivariate analyses, recurrent N3 disease was the only adverse prognostic factor for disease-free and overall survival (P = 0.05 and P = 0.03, respectively). CONCLUSIONS: RND or MRND alone may be the superior treatment for NPC patients with isolated regional failure after primary radiotherapy. Compared to MRND, radical neck dissection could provide better regional control. Postoperative radiotherapy seems to have no benefit on disease-free or overall survival. Distant metastasis is the major cause of death in these patients.

Neck ultrasonography for the evaluation of the etiology of adult unilateral vocal fold paralysis.

BACKGROUND: An extralaryngeal neoplasm involving the vagus nerve and recurrent laryngeal nerve must be excluded when adult unilateral vocal fold paralysis is diagnosed. METHODS: Between 2004 and 2009, 53 adult patients with unilateral vocal fold paralysis received neck ultrasonography. Patients with laryngeal/hypopharyngeal cancer or with previously known cancers of the head and neck, thyroid, esophagus or lung, or with known etiology or trauma were excluded from this study. RESULTS: We included 26 men and 27 women. Ultrasonography revealed 16 patients (30%) having subclinical tumors, including thyroid papillary carcinoma in 7 patients, vagus nerve schwannoma in 2 patients, nodular goiter in 2 patients, malignant nodes in the lower neck in 4 patients, which were metastasized from lung cancer in 3 patients, esophageal cancer in 1 patient, and cervical esophageal cancer in 1 patient. CONCLUSION: Neck ultrasonography is useful to detect subclinical neoplasia, causing unilateral vocal fold paralysis. Thyroid cancer is the most common neoplastic etiology of adult unilateral vocal fold paralysis.

Transcutaneous ultrasound for evaluation of vocal fold movement in patients with thyroid disease.

BACKGROUND: Preoperative evaluation of recurrent laryngeal nerve function is important in the context of thyroid surgery. Transcutaneous ultrasound may be useful to visualize vocal fold movement when evaluating thyroid disease. METHODS: A 7-18 MHz linear array transducer was placed transversely on the midline of the thyroid cartilage at the anterior neck of patients with thyroid disease. The gray-scale technique was used, with the scan setting for the thyroid gland. RESULTS: Between August 2008 and March 2010, 705 patients, including 672 patients with normal vocal fold movement and 33 patients with vocal fold paralysis were enrolled. They included 159 male and 546 female patients. Their ages ranged from 10 to 88 years. Vocal fold movement could be seen by ultrasound in 614 (87%) patients, including 589 (88%) patients with normal vocal fold movement and 25 (76%) patients with vocal fold paralysis (p=0.06). The mean age of patients with visible and invisible vocal fold movement was 46.6 and 57.9 years old, respectively (p=0.001). Ultrasound was able to see vocal fold movement in 533 (98%) female patients but only in 81 (51%) male patients (p=0.001). Among the patients with vocal fold paralysis, ultrasound revealed palsied vocal folds in 17 of 18 (94%) female patients but in only 8 of 15 (53%) male patients (p=0.01). CONCLUSION: Transcutaneous ultrasound represents an alternative tool to evaluate vocal fold movement for more than 85% of patients with thyroid disease, including more than 90% of female patients and about half of male patients.

Feasibility of preservation of the submandibular gland during neck dissection in patients with early-stage oral cancer.

BACKGROUND: The impact of submandibular gland (SMG) preservation during neck dissection on the survival of patients with early-stage oral squamous-cell carcinoma (OSCC) remains undocumented. METHODS: The medical records of all patients with early-stage OSCC (stage I and II) who underwent wide excision of the primary tumor and simultaneous neck dissection between 1999 and 2006 at our facility were retrospectively reviewed. RESULTS: We analyzed 408 patients, including 33 patients with and 375 patients without SMG preservation. The 5-year disease-free and overall survival rates were 78.8% and 90.9% for the patients with SMG preservation and 75.4% and 90.4% for the patients without SMG preservation, and these differences were not statistically significant (P = 0.79, P = 0.99, respectively). Similar survival rates between patients with and without SMG preservation were observed in those with oral tongue squamous-cell carcinoma (SCC) and with buccal SCC. Patients with T2 OSCC with SMG preservation had significantly lower 5-year disease-free survival rate than those without SMG preservation (P = 0.02), but overall survival rates were similar between these two groups. CONCLUSIONS: Preservation of the SMG during neck dissection may be oncologically safe in patients with T1 OSCC, but the feasibility of SMG preservation seems less clear for T2 OSCC.

Postoperative radiotherapy for primary early oral tongue cancer with pathologic N1 neck.

BACKGROUND: The benefit of postoperative radiotherapy (PORT) for early squamous cell carcinoma of the tongue (SCCOT) with pathologic N1 disease remains unclear. METHODS: The medical records of all patients with pathologic T1-2/N0-1 SCCOT who underwent wide excision of the primary tumor and neck dissection between 1980 and 2002 were reviewed. RESULTS: There were 59 patients analyzed, including 28 patients with and 31 patients without PORT. The 5-year disease-free survival rates were 81.2% and 53% for the patients with and without PORT, respectively (p = .03). The overall 5-year survival rates were 77% and 70.5% for the patients with and without PORT, respectively (p = .36). Multivariate analyses showed that PORT had the only protective effect (p = .01) and extracapsular spread (ECS) was the only significantly adverse factor for locoregional recurrence (p = .03). CONCLUSIONS: Approximately one-third of the patients who received only surgery had locoregional recurrence. PORT significantly improved the disease-free survival.

Thyroid cancer incidentally found in radical surgery for laryngeal/hypopharyngeal cancer.

OBJECTIVES: To report on four patients with thyroid cancer found in their specimen of laryngectomy for laryngeal/hypopharyngeal cancer. STUDY DESIGN: Retrospective case series. SETTING: Patients with laryngeal cancer and hypopharyngeal cancers treated at our institution between 1991 and 2005 were enrolled. SUBJECTS AND METHODS: There were 463 patients with laryngeal cancer and 219 patients with hypopharyngeal cancers under retrospective review. RESULTS: There were 254 patients with laryngeal caner and 130 patients with hypopharyngeal cancer receiving surgery. Thyroid cancer was observed in the resected thyroid gland in two cases of laryngeal cancer and in two cases of hypopharyngeal cancer. They received treatment only for their laryngeal/hypopharyngeal cancers. Two patients died of distant metastases from larynx/hypopharynx cancer within one year. The other two patients are currently alive without disease from either of the two types of cancer. CONCLUSIONS: It is rare to discover an unexpected simultaneous thyroid cancer confirmed postoperatively from thyroid tissue partially removed in laryngectomy. A conservative approach is suggested for such patients.

Vocal process granuloma - A result of long-term observation in 53 patients.

Vocal process granulomas are benign lesions involved in the healing of posterior glottis injuries. Here, we report the results of our conservative treatment of vocal process granulomas. Fifty-three patients with 54 occurrences of granulomas between 1998 and 2007 were enrolled. All patients were observed without surgical excision or specific medications and regularly monitored with telescopic examinations until the granulomas disappeared. Data on age, sex, clinical presentation, prior history of intubation or gastroesophageal reflux, telescopic findings, management and clinical course were reviewed. There were 41 males and 12 females. The mean ages of granuloma occurrence were 48.1 years in men and 41.6 years in women (p=0.16). Symptoms included hoarseness in 42 patients, cough in 13 patients, lumping sensation in the throat in 15 patients and sore throat in 14 patients. Ten occurrences were stable during the follow-up period, and the other 44 (81%) achieved complete remission (mean period of 30.6 weeks). The remission patterns included progression and remission in 6 occurrences, slow remission in 22 and rapid remission in 16. The remission rate of granulomas was not significantly related to age (p=0.71), sex (p=0.43), prior intubation (p=0.71), acid reflux (p=0.47), unilateral/bilateral lesions (p=1.00) or granuloma size (p=0.46). The remission time was significantly shorter in patients with prior intubation (p=0.04), but not significantly associated with age (p=0.89), sex (p=0.87), acid reflux (p=0.91), unilateral/bilateral lesions (p=0.26) or granuloma size (p=0.96). Long-term observation has demonstrated that vocal process granulomas can be cured at high remission rates without implementing specific treatments.

[Difficulties facing spouses of in-patients undergoing tocolysis].

When a pregnant woman needs tocolysis to preempt the need for an abortion, there are great impacts on the daily life of her family and on the relationships between the couple and their family members, which negatively influence the balance of the family system. For a pregnant woman under tocolysis in hospital, the spouse indeed plays an important supportive role in helping her to endure many difficulties. Because men are always controllers of power and opinion, being strong and active in attitude to everything, male spouses always suppress their emotions and do not express their feelings to others. These characteristics of men, either in Eastern or Western countries, always cause nursing staff to realize the needs of the male spouse poorly during tocolysis. The current concept of nursing emphasizes family-centered nursing, so the patient's family members are also recipients of treatment, especially male spouses during the process of tocolysis. This is usually neglected, however. In this paper, the author analyzes the characteristics and the roles of male spouses in the family, and the difficulties faced by a pregnant woman during tocolysis in hospital, for the benefit of future clinical nursing practice.

[Study of stress and coping behaviors in families of hospitalized pregnant woman undergoing tocolysis].

Because of tocolysis intervention, families of pregnant woman undergoing tocolysis face stresses beyond the physical and psychological challenges of normal pregnancy. The lifestyle of the family and the relationship between family members will change significantly. The purpose of this study was to explore the stresses and coping behaviors of families of pregnant woman undergoing tocolysis in the hospital. A clinical field method was used and the subjects were recruited from one medical center in Taipei. Data, collected from participant observations and interviews, was subject to content analysis. Results showed that the family stress could be segmented into the four realms of (1) uncertainty about tocolysis; (2) worrying about infant's health; (3) role conflicts between relatives; and (4) changes in family life patterns. The study classified family coping behavior into the following four activity categories: (1) searching for information about tocolysis; (2) adjusting family member roles; (3) changing family member interactions; (4) streamlining the use of family resources. These conclusions are helpful to promote nursing quality in relation to caring for families of pregnant woman with tocolysis by fully understanding the related sources of stress and patterns of coping behaviors.