Factors influencing extrathyroidal extension of papillary thyroid cancer and evaluation of ultrasonography for its diagnosis: a retrospective analysis.

Pathologists usually explore extrathyroidal extensions (ETEs) in thyroid cancer; however, sonographers are often not concerned with ETEs. We investigated factors influencing ETEs and the efficacy of ultrasound evaluation of thyroid capsule invasion. We retrospectively analysed 1933 papillary thyroid carcinoma patients who underwent thyroidectomy during 2018-2021. Patients were divided into three groups: no ETE, minor ETE (mETE), and gross ETE. Clinical characteristic differences were assessed using binary logistic regression analysis to identify ETE predictors, and the kappa test was performed to analyse consistency between ultrasonographic and pathological diagnoses of ETE. The mETE group was more likely to have larger tumour diameters and more extensive lymph node metastasis (LNM) than the no ETE group and more likely to be diagnosed in the isthmus. In the multivariate logistic regression analysis, longest tumour diameter, lesion site, LNM extent, and thyroglobulin concentration were significant mETE predictors. Minimal consistency existed between pathological and ultrasonographic examinations for neighbouring tissue invasion. Many clinical differences were observed between the no ETE and mETE groups, suggesting the importance of considering mETE. Therefore, sonographers should pay more attention to relationships between nodules and capsule and indicate these on ultrasound reports to provide more accurate preoperative ETE information for surgeons.

Increased soluble endoglin levels in newly-diagnosed type 2 diabetic patients are associated with endothelial dysfunction.

Endothelial dysfunction (ED) contributes to the pathologic process underlying macrovascular complications, a common complication of type 2 diabetes mellitus (T2DM). Soluble endoglin (sEng) shed from the extracellular domain of the entire endoglin molecule blocks endothelial protection mediated by transforming growth factor-beta 1 (TGF-ß1). The reactive hyperemia index (RHI), which is determined by reactive hyperemia peripheral arterial tonometry (RH-PAT), is a new index with which to evaluate ED. This study determined the changes in serum sEng levels in newly-diagnosed (untreated) T2DM patients and the correlation with the RHI. The T2DM group included 34 newly-diagnosed T2DM patients, while the control group included 53 healthy adults. The clinical data from the two groups were evaluated retrospectively. The intima-media thickness (IMT) of the common carotid artery (CCA) and the ankle-brachial index (ABI) of both legs were used to assess structural vascular changes. The serum sEng level was determined using an ELISA kit. Endothelial function was assessed using RH-PAT and the RHI was computed. The serum sEng level in the T2DM group was significantly greater than the control group, although the RHI was significantly lower in the T2DM group (p < 0.05). The serum sEng level was negatively correlated with the RHI in T2DM patents (r = 0.354, p = 0.041). The serum sEng level, CCA-IMT, and ABI were not significantly correlated with T2DM (p > 0.05). In summary, among newly-diagnosed T2DM patients, the serum sEng levels were inversely correlated with the RHI, and an elevated sEng level may be associated with ED.

Clinical Value of Ultrasonography and Serum Markers in Preoperative N Staging of Thyroid Cancer.

We aimed to determine factors influencing lymph node metastasis (LNM) and develop a more effective method to assess preoperative N staging. Overall, data of 2130 patients who underwent thyroidectomy for thyroid cancer between 2018 and 2021 were retrospectively analysed. Patients were divided into groups according to pN0, pN1a, and pN1b stages. Pathology was used to analyse the correlation between preoperative serum marker indicators and LNM. Receiver operating characteristic curves were used to compare the diagnostic value of ultrasound (US) examination alone, serum thyroglobulin, age, and combined method for LNM. A significant moderate agreement was observed between preoperative US and postoperative pathology for N staging. Between the pN0 and pN1 (pN1a + pN1b) groups, the differences in free triiodothyronine, anti-thyroid peroxidase antibody, and serum thyroglobulin levels were statistically significant. Among the indicators, serum thyroglobulin was an independent predictor of LNM. The area under the receiver operating characteristic curve was 0.610 for serum thyroglobulin level for predicting LNM, 0.689 for US alone, and 0.742 for the combined method. Both preoperative US and serum thyroglobulin level provide a specific value when evaluating the N staging of thyroid cancer, and the combined method is more valuable in the diagnosis of LNM than US alone.

Effects of Inhaled Corticosteroids on Lung Function in Children With Post-infectious Bronchiolitis Obliterans in Remission.

Background: Post-infectious bronchiolitis obliterans (PIBO) is a rare and irreversible chronic obstructive pulmonary disease with no specific treatment, especially for patients with PIBO in remission. In this study, we evaluated the effects of continuous inhaled corticosteroids (ICSs) and intermittent ICSs on lung function in the remission of PIBO. Methods: This was a retrospective study, and all the subjects we included were divided into continuous ICS group and intermittent ICS group according to treatment regimens. Patients in continuous ICS group received continuous ICSs (2 times a day), and patients in intermittent ICS group received intermittent ICSs (inhaled corticosteroids after acute respiratory tract infection or wheezing). Different lung function tests were performed at different ages. Tidal breathing lung function tests were performed in patients with PIBO aged ≤ 5 years, and the lung ventilation function test and the bronchial dilation test were performed in patients with PIBO aged more than 5 years. Lung function was assessed at the beginning of follow-up and at the end of follow-up (1 year of ICSs). Results: After 1 year of ICSs, patients aged more than 5 years, forced vital capacity (FVC), and forced expiratory volume in 1 s (FEV1) were significantly higher than at the beginning of follow-up. After 1 year of ICSs, the difference in VT/Kg, TPTEF/TE, and VPEF/VE between the end and the beginning of follow-up in continuous ICS group shows an upward trend. But those showed a downward trend in intermittent ICS group. FVC, FEV1, and maximal mid-expiratory flow velocity 25-75% (MMEF25-75%) of continuous ICS group were significantly higher than at the beginning of follow-up. The difference in FEV1 and MMEF25-75% between the end of follow-up and the beginning of follow-up in continuous ICS group was significantly higher than that in intermittent ICS group. A total of 52.94% of patients with PIBO aged more than 5 years were positive for bronchial dilation tests. Conclusion: Inhaled corticosteroids can effectively improve lung function and relieve airway obstruction in patients aged more than 5 years in PIBO remission, especially continuous ICSs. Patients with PIBO may have reversible airflow limitations.

Cytochrome P450 3A4 suppression by epimedium and active compound kaempferol leads to synergistic anti-inflammatory effect with corticosteroid.

Introduction: Cytochrome P450 (CYP) 3A4 is a major drug metabolizing enzyme for corticosteroids (CS). Epimedium has been used for asthma and variety of inflammatory conditions with or without CS. It is unknown whether epimedium has an effect on CYP 3A4 and how it interacts with CS. We sought to determine the effects of epimedium on CYP3A4 and whether it affects the anti-inflammatory function of CS and identify the active compound responsible for this effect. Methods: The effect of epimedium on CYP3A4 activity was evaluated using the Vivid CYP high-throughput screening kit. CYP3A4 mRNA expression was determined in human hepatocyte carcinoma (HepG2) cells with or without epimedium, dexamethasone, rifampin, and ketoconazole. TNF-α levels were determined following co-culture of epimedium with dexamethasone in a murine macrophage cell line (Raw 264.7). Active compound (s) derived from epimedium were tested on IL-8 and TNF-α production with or without corticosteroid, on CYP3A4 function and binding affinity. Results: Epimedium inhibited CYP3A4 activity in a dose-dependent manner. Dexamethasone enhanced the expression of CYP3A4 mRNA, while epimedium inhibited the expression of CYP3A4 mRNA and further suppressed dexamethasone enhancement of CYP3A4 mRNA expression in HepG2 cells (p < 0.05). Epimedium and dexamethasone synergistically suppressed TNF-α production by RAW cells (p < 0.001). Eleven epimedium compounds were screened by TCMSP. Among the compounds identified and tested only kaempferol significantly inhibited IL-8 production in a dose dependent manner without any cell cytotoxicity (p < 0.01). Kaempferol in combination with dexamethasone showed complete elimination of TNF-α production (p < 0.001). Furthermore, kaempferol showed a dose dependent inhibition of CYP3A4 activity. Computer docking analysis showed that kaempferol significantly inhibited the catalytic activity of CYP3A4 with a binding affinity of -44.73kJ/mol. Discussion: Inhibition of CYP3A4 function by epimedium and its active compound kaempferol leads to enhancement of CS anti-inflammatory effect.

Establishment of a predictive equation for pulmonary ventilation function in school-aged children in northeast China: a prospective study. / ä¸œåŒ—åœ°åŒºå­¦é¾„æœŸå„¿ç«¥è‚ºé€šæ°”åŠŸèƒ½é¢„è®¡æ–¹ç¨‹å¼å»ºç«‹çš„å‰çž»æ€§ç ”ç©¶.

OBJECTIVES: To establish a predictive equation for commonly used pulmonary ventilation function parameters in children aged 6-<16 years in northeast China. METHODS: A total of 504 healthy children from Liaoning, Jilin, and Heilongjiang provinces of China were selected for the prospective study, among whom there were 242 boys and 262 girls. The JAEGER MasterScreen Pneumo spirometer was used to measure pulmonary ventilation function. With the measured values of 10 parameters, including forced vital capacity (FVC), forced expiratory volume in 1 second (FEV1), FEV1/FVC ratio, and back-extrapolated volume (BEV), as dependent variables and age, body height, and body weight as independent variables, the stepwise multivariate regression method was used to establish the regression equation for children of different sexes. The mean of relative prediction error was used to evaluate the applicability of the predictive equation. RESULTS: The boys aged 9-<10 years and 15-<16 years had significantly higher body height, FVC, and FEV1 than the girls of the same age (P<0.05), and the boys aged 9-<10, 10-<11, 11-<12, and 13-<14 years had a significantly lower FEV1/FVC ratio than the girls of the same age (P<0.05). The correlation analysis showed that all parameters, except FEV1/FVC ratio and BEV/FVC ratio, were significantly positively correlated with age, body height, and body weight (P<0.001). Further regression analysis showed that age and body height were the influencing factors for most parameters, while body weight was less frequently included in the regression equation. Compared with the predictive equations from previous studies, the regression equation established in this study had relatively good applicability in the study population. CONCLUSIONS: A new predictive equation for the main pulmonary ventilation function parameters has been established in this study for children aged 6-<16 years in northeast China, which provides a basis for accurate judgment of pulmonary function abnormalities in clinical practice.

Insight Into an Outbreak of Canine Distemper Virus Infection in Masked Palm Civets in China.

In August 2019, a suspected outbreak of canine distemper was observed in a masked palm civet farm that also received stray civets and rescued wild civets in Henan Province of China. A virulent canine distemper virus (CDV) strain, named HN19, from vaccinated masked palm civets was the etiologic agent identified in this outbreak using RT-PCR and sequencing of the complete genome. Serological analysis indicated a lower positive rate of CDV-neutralizing antibody in wild civets than in captive civets. Phylogenetic analysis of viral hemagglutinin (H) and the complete genome showed high identities with Rockborn-like strains at the nucleotide (98.7~99.72%) and the closest nucleotide similarity with a strain that killed lesser pandas in China in 1997, but low identities with America-1 strains (vaccine strains). Most importantly, one distinct amino acid exchange in the H protein at position 540 Asp → Gly (D540G), which confers CDV with an improved ability to adapt and utilize the human receptor, was observed in HN19. This study represents the first reported outbreak of a Rockborn-like CDV strain infection in masked palm civets in China. Based on this report, the existence of Rockborn-like strains in Chinese wild animals may not only cause immune failure in captive animals, but may also confer increased zoonotic potential.

Epidemiology of Adenovirus Pneumonia and Risk Factors for Bronchiolitis Obliterans in Children During an Outbreak in Jilin, China.

Background: Jilin Province, located in northeastern China, recently experienced a human adenovirus (HAdV) epidemic. Few studies involving hospitalized pediatric patients with pneumonia caused by HAdV in our region exist. HAdV pneumonia can lead to severe long-term respiratory sequelae, such as post-infectious bronchiolitis obliterans (PIBO), which has a poor prognosis and greatly influences the quality of life of pediatric patients. However, studies on the risk factors for PIBO are limited. Objective: To describe the HAdV pneumonia prevalence and determine potential risk factors for PIBO development among hospitalized children in Jilin Province, China. Methods: The data of 187 children with HAdV pneumonia (10 months-12 years old) admitted to the First Hospital of Jilin University during an outbreak between October 2018 and January 2020 were retrospectively studied. We analyzed the epidemiological characteristics of HAdV pneumonia, focusing on severe HAdV pneumonia (66 cases). The risk factors for BO development were determined by comparing the demographic and clinical data of the BO and non-BO groups. Results: The largest number of HAdV pneumonia cases occurred in January 2019 (severe n = 18, general n = 21), followed by December 2018 (severe n = 14, general n = 11), June 2019 (general n = 17), July 2019 (general, n = 14), and May 2019 (general, n = 13). In total, 91.98% of the children with HAdV pneumonia were <6 years old (172/187), and 50% of the pediatric patients with severe HAdV pneumonia were <2 years old (33/66). We found that 30.3% of the severe cohort developed BO (20/66), and the strongest independent risk factors for PIBO were persistent wheezing (OR 181.776, 95% CI, 3.385-9,761.543) and acute respiratory failure (OR 51.288, 95% CI, 1.858-1,415.441) during a severe pneumonia episode. Conclusions: The largest number of HAdV pneumonia cases, especially severe cases, occurred in winter in Northeast China, followed by summer. The majority of children admitted with HAdV pneumonia were <6 years old, and half of severe HAdV pneumonia patients were <2 years old. Children who had persistent wheezing or acute respiratory failure during the acute phase of severe HAdV pneumonia were prone to the development of BO.

Longitudinal Assessment of Pulmonary Function and Bronchodilator Response in Pediatric Patients With Post-infectious Bronchiolitis Obliterans.

Backgroud: Postinfectious bronchiolitis obliterans (PIBO) is a rare respiratory disease. In recent years, the disease has been recognized and diagnosed increasingly in children. Pulmonary function is important for diagnosis, identifying the severity of the PIBO and monitoring progression. But there have been only a few studies that followed the evolution of PIBO on the basis of pulmonary function tests (PFTs). Objective: The study targeted the evolution of pulmonary function and bronchodilator response in a case series of Chinese children with PIBO. Methods: Twelve children between the ages of 6-99 months with PIBO were studied retrospectively from 2009 to 2019. Forced vital capacity (FVC), forced expiratory volume in 1 s (FEV1), the FEV1/FVC ratio, and maximal midexpiratory flow velocity 25-75% (MMEF25-75%) were collected at each PFT, and bronchodilator responses were evaluated. Spirometric parameters were monitored over time, and generalized linear mixed models were used to analyze longitudinal panel data. Results: The median baseline PFT values for FVC, FEV1, the FEV1/FVC ratio, and MMEF25-75% were 41.6, 39.75, 90.7, and 22.2%, respectively. At the initial PFTs, 10 (83.3%) patients demonstrated a significant bronchodilator response. FVC and FEV1 increased by 8.212%/year and 5.007%/year, respectively, and the FEV1/FVC ratio decreased by an average of 3.537%/year. MMEF25-75% showed improvement at an average rate of 1.583% every year. Overall, FEV1 and MMEF25-75% showed different degrees of improvement after the use of inhaled bronchodilators at each PFT session for 10 patients, and FEV1 measures demonstrated significant (>12%) ß2-bronchodilation in 56% of PFT sessions. Conclusions: Pediatric patients with PIBO showed an obstructive defect in pulmonary function. The FVC, FEV1, and MMEF25-75% improved as they grew older, while the FEV1/FVC ratio decreased. This may be due to the development of lung parenchyma more than airway growth. Airway obstruction in some patients improved with the use of ß2 agonists.

CircRNA_0050463 promotes influenza A virus replication by sponging miR-33b-5p to regulate EEF1A1.

Circular RNAs (circRNAs), a new class of widely expressed endogenous regulatory RNAs, are characterized by a covalently closed loop structure without a 5' cap or 3' tail. Increasing numbers of studies have shown that circRNAs play important roles in diverse physiological and pathological processes, including the dynamic interactions between viruses and hosts. However, their multifaceted roles in cellular responses to influenza A virus (IAV) infection remain largely unknown. Here, we analyzed the expression of circ_0050463, which is predominantly localized in cytoplasm, in response to IAV infection. Knockdown of circ_0050463 with siRNA in A549 cells inhibited IAV replication. In addition, the activation of nuclear factor κB (NF-κB) was involved in IAV-induced circ_0050463 expression, as revealed by assay using a NF-Kb inhibitor (Bay 11-7082). By performing biotin-labeled RNA pull-down and luciferase reporter assay, we demonstrated that circ_0050463 functioned as an endogenous microRNA-33b-5p sponge to sequester and inhibit miR-33b-5p activity, resulting in increased eukaryotic translation elongation factor 1 alpha 1 (EEF1A1) expression with subsequent facilitation of IAV replication. Taken together, the results of our study indicate that the circ_0050463 promotes IAV replication via miR-33b-5p/EEF1A1 axis, thus providing evidence for the host circRNAs utilized by viruses to support their replication.

RET S409Y Germline Mutation and Associated Medullary Thyroid Carcinoma.

Background: Inherited medullary thyroid carcinoma (MTC) is primarily caused by RET mutations that are commonly localized in exons 5, 8, 10, 11, and 13-16. In this study, we report pedigrees for individuals with MTC that harbor a germline S409Y variant within exon 6 of the RET proto-oncogene. Methods: Targeted sequencing was used to diagnose four apparently sporadic MTC index cases carrying the germline RET S409Y (c.1226 C>A) variant. Subsequently, 27 relatives of these individuals underwent clinical and genetic assessments and/or thyroid surgery. Furthermore, in silico analyses and in vitro assays were performed to predict or verify the potential oncogenic activity of the S409Y variant. Results: Overall, 15 of 31 participants were found to carry the RET S409Y variant. Of these, 6 presented with isolated MTC (mean age 50.2 years; range 41-75 years), of which 3 presented with neck lymph node metastases and 2 presented with distant liver or lung metastases. Among the remaining 9 carriers, 3 (mean age 56 years; range 41-76 years) had elevated serum calcium-stimulated calcitonin (sCtn) or concurrent marginally elevated serum calcitonin (Ctn) levels, whereas the other 6 (mean age 37.5 years; range 14-52 years) exhibited typical Ctn/sCtn levels (p < 0.05). None of the 15 carriers in these 4 families presented clinical evidence of pheochromocytoma, hyperparathyroidism, or Hirschsprung's disease. In silico analyses revealed that S409Y was a "possibly damaging" mutation that could affect the RET protein inter-domain interface. An in vitro assay revealed that the phosphorylation level of RET tyrosine 905 was relatively higher in the RET S409Y mutant than in wild-type (WT) RET. Moreover, transfection of HEK 293 cells with S409Y enhanced the phosphorylation activity of AKT, ERK pathways, and it increased cell proliferation compared with WT RET, but to a lesser degree than that for the RET C618Y and C634Y mutations. Conclusions: This study demonstrates that the novel germline RET S409Y variant is likely pathogenic and is associated with lower penetrance of MTC than that for the C618Y and C634Y mutations. Individuals with S409Y should be managed using a personalized approach, and additionally, "at-risk" family members should be evaluated. Additional studies are needed to elucidate the correlation between the S409Y mutation and multiple endocrine neoplasia type 2-specific tumors.

A case of Kawasaki disease presenting with parotitis: A case report and literature review.

RATIONALE: Kawasaki disease affects multiple organ systems. Its typical symptoms include fever, rash, oropharyngeal mucosal erythema, bilateral non-exudative conjunctivitis, cervical lymphadenopathy, extremity changes, and membranous desquamation of the fingers and toes. In severe cases, cardiovascular, respiratory, musculoskeletal, gastrointestinal, neurological, and genitourinary complications may occur. In the early stage, Kawasaki disease is often manifested by uncommon symptoms, such as pyuria, meningitis, shock, and retropharyngeal or parapharyngeal abscess, which may delay diagnosis and treatment. We have reported a case of Kawasaki disease presenting with mumps and reviewed the clinical features of 14 other similar cases, in order to facilitate the early diagnosis and treatment of this unusual presentation of Kawasaki disease. PATIENT CONCERNS: A 10-year-old boy presented with persistent fever and parotitis and was diagnosed with suppurative parotitis. After antibiotic therapy, the parotid swelling reduced, but the fever persisted and other typical symptoms of Kawasaki disease appeared, including bilateral conjunctival hyperaemia, cervical lymphadenopathy, oropharyngeal mucosal erythema, membranous desquamation of the fingers, and left coronary artery widening. DIAGNOSES: The patient was diagnosed with Kawasaki disease 12 days after the onset of fever. INTERVENTIONS: The patient was administered γ-globulin 1.0 g/kg·d for 2 consecutive days and oral aspirin 5 mg/kg·d. OUTCOMES: The left coronary artery returned to a width of 3.8 mm after 1 month and of 3.1 mm after 3 months. The dose of aspirin was reduced to 3 mg/kg·d after 2 months and to 1.5 mg/kg·d after 3 months. LESSONS: Physicians should be aware that Kawasaki disease may develop after parotitis.

IL-23 and dendritic cells: What are the roles of their mutual attachment in immune response and immunotherapy?

Interleukin-23 (IL-23) is a cytokine that is composed of the subunits p19 and p40, while its receptor (IL-23R) consists of two subunits, that is, IL-23Rα and IL-12Rß1. The interaction between IL-23 and IL-23R is necessary for exerting cardinal biological effects upon certain cell types, including promotion of memory T cell proliferation and Th17 cell-mediated IL-17 secretion. Accordingly, dendritic cells (DCs) are one of the main sources for IL-23 secretion. Interestingly, IL-23R is also present on the DC plasma membrane, suggesting that IL-23 potentially acts on DCs via an autocrine manner. In this review, we have summarized a variety of IL-23-mediated effects on the intracellular signaling pathways such as Janus kinase 2, tyrosine kinase 2, signal transducer and activator of transcription (STAT), mitogen-activated protein kinase signaling, and so forth, which may underlie numerous processes such as DC maturation, antigen presentation, T cell proliferation/activation, and cytokine secretion, which may be implicated in many immune-related diseases through IL-23/DC interactions. Accordingly, these signaling pathways are extensively involved in the pathogenesis and progression of numerous diseases, including autoimmune disease (e.g., atopic dermatitis, asthma, and multiple sclerosis) and infection (e.g., bacterial, fungal, and viral infections). Taken together, they are potentially applicable to novel but promising strategies for treating numerous diseases associated with the mutual attachment of IL-23 and DCs.

Requirement of TGFß Signaling for Effect of Fluoride on Osteoblastic Differentiation.

Research focused on transforming growth factor ß (TGFß) signaling in osteoblast is gradually increasing, whereas literature is rare in terms of fluorosis. This work aimed to investigate how TGFß signaling participated in regulation of the osteoblast by different doses of fluoride treatment. Bone marrow stem cells (BMSCs) were developed into osteoblastic cells and exposed to 1, 4, and 16 mg/L F- with and without 10 ng/mL of TGFß. Cell viability and differentiation state of osteoblast under different settings were measured by means of cell counting kit and analysis of alkaline phosphatase (ALP) activity as well as formation of mineral nodules. Real-time PCR was utilized to test expression of ALP and Runt-related transcription factor 2 (Runx2) at gene level. The gene expression of TGFß signaling effectors was also investigated, such as TGFß receptors (TßRs), smad3, and mitogen-activated protein kinases (MAPK). Results demonstrated that fluoride treatment exhibited action on osteoblast viability and osteogenic differentiation and upregulated expression of TßR2, smad3, and MAPK in this process. Administration of TGFß strengthened ALP activity but attenuated formation of mineral nodules. Co-treatment of TGFß and low-dose fluoride increased ALP activity compared to same dose of single fluoride treatment, whereas it inhibited mineral nodule formation. Administration of TGFß reversed the suppression of high-dose fluoride on osteogenic differentiation of BMSCs. Taken together, studies revealed that TßR2 acted as a target for fluoride and TGFß treatment on BMSCs, and smad3 and MAPK were involved in the mechanism of fluoride regulating osteogenic differentiation. Together, our data indicated that TGFß receptor-mediated signaling through smad3 and MAPK was required for modulation of fluoride on osteoblast viability and differentiation, and activating TßR2-smad3 signaling pathway reversed suppression of osteoblasts differentiation by high dose of fluoride treatment.

The RET C611Y mutation causes MEN 2A and associated cutaneous

Background: Cutaneous lichen amyloidosis (CLA) has been reported in some multiple endocrine neoplasia type 2A (MEN 2A) families affected by specific germline RET mutations C634F/G/R/W/Y or V804M, as a characteristic of the clinical manifestation in 'MEN 2A with CLA', one of four variants of MEN 2A, which was strictly located in the scapular region of the upper back. Patient Findings: This study reports a large south-eastern Chinese pedigree with 17 individuals carrying the MEN 2A-harboring germline C611Y (c.1832G>A) RET mutation by Sanger sequencing. One individual presented MEN 2A-related clinical features, including typical CLA in the interscapular region; another individual exhibited neurological pruritus and scratching in the upper back but lacked CLA skin lesions. Both subjects presented with CLA or pruritic symptoms several years before the onset of medullary thyroid carcinoma (MTC) and/or pheochromocytoma. The remaining 15 RET mutation carriers did not exhibit CLA; of these, one presented with MTC and pheochromocytoma, nine with MTC only, two with elevated serum calcitonin and three younger subjects with normal serum calcitonin levels. This family's clinical data revealed a later diagnosis of MTC (mean age, 45.9 (range: 23­73) years), a lower penetrance of pheochromocytoma (2/17, 11.8%) and CLA (1/17, 5.9%). However, no hyperparathyroidism and Hirschsprung disease were reported in this family. Summary and Conclusions: This is the first description of a family with MEN 2A-related CLA due to a germline RET C611Y mutation, which might exhibit a novel and diversified genotype­phenotype spectrum in MEN 2A.

PTH (1-34) affects bone turnover governed by osteocytes exposed to fluoride.

Exposure to fluoride from environmental sources remains an overlooked, but serious public health risk. In this study, we looked into the role osteocytes play on the mechanism underlying fluoride induced osteopathology. We analyzed bone formation and resorption related genes generated by osteocytes that were exposed to varied doses of fluoride with and without PTH in vitro. Correspondingly, osteogenesis and osteoclastogenesis related genes were also investigated in rats exposed to fluoride for 8 weeks, and the PTH(1-34)was applied at the last 3 weeks to observe its role in regulating bone turnover upon fluoride treatment. The data in vitro indicated that fluoride treatment inhibited Sost expression of mRNA and protein and stimulated RANKL mRNA protein expression as well as the RANKL/OPG ratio in the primary osteocytes. Single PTH treatment played the similar role on expression of these genes and proteins. The PTH combined administration enhanced the action of fluoride treatment on RNAKL/OPG and SOST/Sclerostin. The up-regulation of RANKL and decreasing of Sost induced by fluoride and/or PTH treatment was validated in vivo and suggests that osteocytes are a major source of RANKL and Sost, both of which play essential roles in fluoride affecting osteogenesis and osteoclastogenesis. Expression of Wnt/ß-catenin was up-regulated in both in vitro osteocytes treated with high dose of fluoride and bone tissue of rats in the presence of fluoride and PTH. In vivo, fluoride and single PTH stimulated bone turnover respectively, furthermore, PTH combined with low dose of fluoride treatment reinforced the osteogenesis and osteoclastogenesis genes expression, however, co-treatment of PTH reversed the effect of high dose of fluoride on osteogenesis and osteoclastogenensis related factors. In conclusion, this study demonstrated that osteocytes play a key role in fluoride activated bone turnover, and PTH participates in the process of fluoride modulating SOST/Sclerostin and RANKL expression.

The role of TGFß receptor 1-smad3 signaling in regulating the osteoclastic mode affected by fluoride.

Studies that have focused on the role TGFß signaling plays in osteoclast activity are gradually increasing; however, literature is rare in terms of fluorosis. The aim of this study is to observe the role the TßR1/Smad3 pathway plays in fluoride regulating cellsosteoclast-like cells that are under the treatment of TGFß receptor 1 kinase. The RANKL-mediated osteoclast-like cells from RAW264.7 cells were used as osteoclast precursor model. The profile of miRNA expression in fluoride-treated osteoclast-like cells exhibited 303 upregulated miRNAs, 61 downregulated miRNAs, and further drew 37 signaling pathway maps by KEGG and Biocarta pathway enrichment analysis. TGFß and its downstream effectors were included among them. Osteoclast viability, formation and function were detected via MTT method, bone resorption pit and tartrate-resistant acid phosphatase (TRACP) staining, respectively. Results demonstrated that different doses of fluoride exhibited a biphasic effect on osteoclast cell viability, differentiation, formation and function. It indicated that a low dose of fluoride treatment stimulated them, but high dose inhibited them. SB431542 acted as TßR1 kinase inhibitor and blocked viability, formation and function of osteoclast-like cells regulated by fluoride. The expression of the osteoclast marker, RANK, and TßR1/Smad3 at gene and protein level was analyzed under fluoride with and without SB431542 treatment. Fluoride treatment indicated little effect on the RANK protein expression; however it significantly influenced TRACP expression in osteoclast-like cells. The stimulation of fluoride on the expression of Smad3 gene and phosphorylated Smad3 protein exhibited dose-dependent manner. SB431542 significantly impeded phosphorylation of Smad3 protein and TRACP expression in osteoclast-like cells that were exposed to fluoride. Our work demonstrated that TGFß signaling played a key role in fluoride regulating osteoclast differentiation, formation and function. It elucidated that TßR1/Smad3 pathway participated in the mechanism of biphasic modulation of osteoclast mode regulated by fluoride.

Immunotherapeutic effects of cytokine-induced killer cells combined with CCL21/IL15 armed oncolytic adenovirus in TERT-positive tumor cells.

The effective antitumor immune responses are dependent on coordinate interaction of various effector cells. Thus, the combination of adoptive immunotherapy and target gene therapy is capable of efficiently generating a productive antitumor immune response. We investigated whether combination of cytokine-induced killer (CIK) cells adoptive immunotherapy and CCL21/IL15 armed oncolytic adenovirus could induce the enhanced antitumor activity. The CCL21/IL15 co-expression oncolytic adenoviruses were constructed by using the AdEasy system, which uses homologous recombination with shuttle plasmids and full length Ad backbones. This conditionally replicating adenoviruses CRAd-CCL21-IL15 could induce apoptosis in TERTp-positive tumor cells for viral propagation, but do not replicate efficiently in normal cells, because the E1A promoter was replaced by telomerase reverse transcriptase promoter (TERTp). Our results showed that the combination of CIK cells and CRAd-CCL21-IL15 could induce higher antitumor activity than either CIK cells or CRAd-CCL21-IL15 alone. This combined treatment could induce the tumor specific cytotoxicity of CTLs (cytotoxic T lymphocytes) in vitro. Moreover, the treatment of established tumors with the combined therapy of CIK cells and CRAd-CCL21-IL15 resulted in tumor regression. This study suggests that the combined treatment by adoptive immunotherapy and gene therapy is a promising strategy for the therapy of tumor.

Analysis of the Role of Insulin Signaling in Bone Turnover Induced by Fluoride.

The role of insulin signaling on the mechanism underlying fluoride induced osteopathology was studied. We analyzed the expression of genes related with bone turnover and insulin signaling in rats treated by varying dose of fluoride with or without streptozotocin (STZ) in vivo. Furthermore, insulin receptor (InR) expression in MC3T3-E1 cells (pre-osteoblast cell line) was interfered with small interfering RNA (siRNA), and genes related with osteoblastic and osteoclastic differentiation were investigated in cells exposed to fluoride in vitro for 2 days. The in vivo study indicated the possible role of insulin in bone lesion induced by excessive amount of fluoride. Fluoride activated the InR and Insulin-like growth factor 1 (IGF1) signaling, which were involved in the mechanism underlying fluoride induced bone turnover. The TGFß1 and Wnt10/ß-catenin pathway took part in the mechanism of bone lesion induced by fluoride, and insulin probably modulated the TGFß1 and ß-catenin to exert action on bone turnover during the development of bone lesion. The in vitro study showed the concomitant decrease of OPG, osterix and OCN with inhibition of InR expression in osteoblast, and three genes still was low in cells co-treated with fluoride and InR siRNA, which suggested that fluoride probably stimulated the expression of OPG, osterix and OCN through InR signaling. In conclusion, insulin played the important role in bone lesion induced by excessive amount of fluoride through mediating InR receptor signaling, and IGF1 signaling probably exerted action on bone turnover caused by overdose of fluoride.

Structural identification of neopanaxadiol metabolites in rats by ultraperformance liquid chromatography/quadrupole-time-of-flight mass spectrometry.

RATIONALE: Neopanaxadiol (NPD) is one of the major ginsenosides in Panax ginseng C. A. Meyer (Araliaceae) that has been suggested to be a drug candidate against Alzheimer's disease. However, few data are available regarding its metabolism in rats. METHODS: In this study, a method of ultraperformance liquid chromatography/quadrupole-time-of-flight mass spectrometry (UPLC/QTOFMS) was developed to identify major metabolites of NPD in the stomach, intestine, urine and feces of rats, with the aim of determining the main metabolic pathways of NPD in rats after oral administration. RESULTS: UPLC/QTOFMS revealed two metabolites in the stomach of rats, one metabolite in the intestine and two metabolites in feces. One metabolite, named M2, was isolated and purified from rats feces, which was identified as (20S,22S)-dammar-22,25-epoxy-3ß,12ß,20-triol based on extensive NMR spectroscopy and mass spectrometry data. The main metabolites of NPD in rats were the products of epoxidation, dehydrogenation and hydroxylation. NPD was predominantly metabolized by 20,22-double-bond epoxidation and rearrangement to yield an expoxidation product (M2). CONCLUSIONS: Based on the profiles of the metabolites, possible metabolic pathways of NPD in rats were proposed for the first time. This study provides new and available information on the metabolism of NPD, which is indispensable for further research on metabolic pathways of dammarane ginsengenins in vivo.