Phylogenetic relationships of Grapsoidea and insights into the higher phylogeny of Brachyuran.

Decapoda is one of the most diverse crustacean orders, and has become an important research subject. However, the phylogenetic relationships among the main lineages of Decapoda remain uncertain, especially in the order Brachyura. Herein, we sequenced the whole mitochondrial genome of V. litterata and constructed a phylogenetic tree to understand its phylogenetic relationships with other species. The results showed that the mitochondrial genome of V. litterata was generally similar to mitogenomes of Metazoa reported in the literature, with a size of 16,247 bp, 37 genes, and a control region. Both AT-skew and GC-skew were negative, indicating more abundant Cs and Ts than Gs and As. The gene arrangement of V. litterata is identical to those of Eriocheir hepuensis, Cyclograpsus granulosus, Hemigrapsus sanguineus, Helicana wuana, and Helice tientsinensis but differs from the pancrustacean ground pattern and typical arrangement of Brachyuran crabs. Phylogenetic reconstruction showed that V. litterata belongs to the Varunidae.

Peroxiredoxin 6 modulates Toll signaling pathway and protects DNA damage against oxidative stress in red swamp crayfish (Procambarus clarkii).

Peroxiredoxin 6 (Prx6) is an important member of the peroxiredoxin family that plays critical roles in protecting host against the toxicity of oxidative stress and participates in cell signaling. Herein, we report Prx6 gene from red swamp crayfish, Procambarus clarkii. The cDNA fragment of PcPrx6 was 660 bp, encoding a 219 amino acid residues protein. The quantitative real time PCR analysis showed ubiquitous expression of PcPrx6 mRNA in the tested tissues. The challenge with peptidoglycan and Poly I:C remarkably suppressed the mRNA level of PcPrx6 in hepatopancreas at 3, 12, 48 h compared with the PBS control. However, the expression level significantly increased after 36 h of their treatment. The knockdown of PcPrx6 by small interference RNA significantly enhanced the transcript levels of Toll pathway-responsive genes at 24 h. Recombinant PcPrx6 protein was purified using affinity chromatography and analyzed for its biological role. The results revealed that the recombinant PcPrx6 protein manifested the ability to protect supercoiled DNA damage from oxidative stress elicited by mixed function oxidative assay. Altogether, PcPrx6 may have multiple functional roles in the physiology of P. clarkii, since it negatively regulates the Toll signaling transduction and protects supercoiled DNA damage from oxidative stress.

Essential role of the peroxiredoxin 4 in Procambarus clarkii antioxidant defense and immune responses.

Peroxiredoxin (Prx) family members play a key role in host defense against oxidative stress, and modulate immune responses following microbial infection. Here, we cloned and characterized Procambarus clarkii Prx4 (Peroxiredoxin 4) cDNA, a regulator of oxidative stress and its expression analysis upon lipopolysaccharide (LPS) and polyriboinosinic polyribocytidylic acid (Poly I:C) infection. The cDNA fragment of PcPrx4 was 744 bp in length, encoding a putative protein of 248 amino acid residues. Real-time quantitative reverse transcription-PCR (qRT-PCR) analysis showed that the PcPrx4 was expressed in all the examined tissues, and it was highest in the hepatopancreas followed by the hemocytes and gill. The challenge with LPS and Poly I:C significantly up-regulated the expression of PcPrx4 in hepatopancreas, hemocytes and gill when compared with the control. Recombinant PcPrx4 protein was used to investigate the antioxidant function in vitro by mixed-function oxidase assay. The results demonstrated a dose-dependent inhibition of DNA damage by rPcPrx4 protein. Altogether, our results imply that PcPrx4 is implicated in defense against microbial pathogens and oxidants in P. clarkii.

Notch3 expression correlates with thyroid cancer differentiation, induces apoptosis, and predicts disease prognosis.

BACKGROUND: Thyroid tumorigenesis is characterized by a progressive loss of differentiation exhibited by a range of disease variants. The Notch receptor family (1-4) regulates developmental progression in both normal and cancerous tissues. This study sought to characterize the third Notch isoform (Notch3) across the various differentiated states of thyroid cancer, and determine its clinical impact. METHODS: Notch3 expression was analyzed in a tissue microarray of normal and pathologic thyroid biopsies from 155 patients. The functional role of Notch3 was then investigated by upregulating its expression in a follicular thyroid cancer (FTC) cell line. RESULTS: Notch3 expression regressed across decreasingly differentiated, increasingly malignant thyroid specimens, correlated with clinicopathological attributes reflecting poor prognosis, and independently predicted survival following univariate and multivariate analyses. Overexpression of the active Notch3 intracellular domain (NICD3) in a gain-of-function FTC line led to functional activation of centromere-binding protein 1, while increasing thyroid-specific gene transcription. NICD3 induction also reduced tumor burden in vivo and initiated the intrinsic apoptotic cascade, alongside suppressing cyclin and B-cell lymphoma 2 family expression. CONCLUSIONS: Loss of Notch3 expression may be fundamental to the process of dedifferentiation that accompanies thyroid oncogenesis. Conversely, activation of Notch3 in thyroid cancer exerts an antiproliferative effect and restores elements of a differentiated phenotype. These findings provide preclinical rationale for evaluating Notch3 as a disease prognosticator and therapeutic target in advanced thyroid cancer. Cancer 2017;123:769-82. © 2016 American Cancer Society.

Generation of Novel Thyroid Cancer Stem-Like Cell Clones: Effects of Resveratrol and Valproic Acid.

Anaplastic thyroid cancer is an aggressive and highly lethal cancer for which conventional therapies have proved ineffective. Cancer stem-like cells (CSCs) represent a small fraction of cells in the cancer that are resistant to chemotherapy and radiation therapy and are responsible for tumor reoccurrence and metastasis. We characterized CSCs in thyroid carcinomas and generated clones of CSC lines. Our study showed that anaplastic thyroid cancers had significantly more CSCs than well-differentiated thyroid cancers. We also showed that Aldefluor-positive cells revealed significantly higher expression of stem cell markers, self-renewal properties, thyrosphere formation, and enhanced tumorigenicity. In vivo passaging of Aldefluor-positive cells resulted in the growth of larger, more aggressive tumors. We isolated and generated two clonal spheroid CSC lines derived from anaplastic thyroid cancer that were even more enriched with stem cell markers and more tumorigenic than the freshly isolated Aldefluor-positive cells. Resveratrol and valproic acid treatment of one of the CSC lines resulted in a significant decrease in stem cell markers, Aldefluor expression, proliferation, and invasiveness, with an increase in apoptosis and thyroid differentiation markers, suggesting that these cell lines may be useful for discovering new adjuvant therapies for aggressive thyroid cancers. For the first time, we have two thyroid CSC lines that will be useful tools for the study of thyroid CSC targeted therapies.

A role of cathepsin L gene in innate immune response of crayfish (Procambarus clarkii).

Cathepsin L is one of the crucial enzyme superfamilies and involved in the immune responses. In the present study, cathepsin L gene from the red crayfish Procambarus clarkii, named PcCTSL, was cloned and characterized. The cDNA fragment of PcCTSL was 1026 bp in length, which encoded a putative protein of 341 amino acid residues with a molecular weight of 37.884 kDa. The theoretical isoelectric point was 5.218. The prepro-cathepsin L was comprised of a typical signal peptide (Met1-Ala18), a prodomain proregion peptide (Trp29-Phe89) and a mature peptide (Leu124-Leu340). Homology analysis indicated that PcCTSL exhibited 53.2%-87.1% identity to other selected species. The recombinant protein of PcCTSL was successfully expressed in Escherichia coli and rabbit anti-PcCTSL polyclonal antibodies were prepared. Real-time quantitative reverse transcription-PCR (qPCR) analysis revealed that the PcCTSL was expressed in all examined tissues, while the greatest mRNA level was observed in hepatopancreas. The expression of PcCTSL mRNA was clearly up regulated in hepatopancreas after challenge by lipopolysaccharide (LPS) and polyriboinosinic polyribocytidylic acid (Poly I:C). RNA interference of PcCTSL affected the gene expression of members of the Toll pathway. Our results suggest that the PcCTSL may play an important role to defend P. clarkii against the pathogens infection.

[Nontargeted metabolomic analysis of Anoectochilus roxburghii at different cultivation stages].

Anoectochilus roxburghii is a traditional Chinese medicine and natural health products. In the modern cultivation system, A. roxburghii is micropropagated in tissue culture, and the plants are transferred to soil cultivation for months. However, it remains unclear about the necessity of soil cultivation for the accumulation of health beneficial compounds. In this paper, we performed nontargeted metabolomic analysis using GC-TOF-MS and UPLC-Q-TOF-MS, on A. roxburghii plants at tissue culture stage or after 3 months of soil cultivation. The results showed that the primary metabolites such as alcohols and organic acids are abundant in the tissue culture plants. In contrast, polysaccharide, nucleoside, esters and secondary metabolites such as flavonoids, terpenoids were significantly accumulated in cultivated seedlings. Flavonoids and polysaccharides are considered as the principle effective components in A. roxburghii. Soil cultivation period is therefore essential for the accumulation of these metabolites.

The roles of the epithelial-mesenchymal transition marker PRRX1 and miR-146b-5p in papillary thyroid carcinoma progression.

Thyroid carcinoma is the most common endocrine malignancy, and papillary thyroid carcinoma represents the most common thyroid cancer. Papillary thyroid carcinomas that invade locally or metastasize are associated with a poor prognosis. We found that, during epithelial-mesenchymal transition (EMT) induced by transforming growth factor-ß1 (TGF-ß1), papillary thyroid carcinoma cells acquired increased cancer stem cell-like features and the transcription factor paired-related homeobox protein 1 (PRRX1; alias PRX-1), a newly identified EMT inducer, was markedly up-regulated. miR-146b-5p was also transiently up-regulated during EMT, and in siRNA experiments miR-146b-5p had an inhibitory role on cell proliferation and invasion during TGF-ß1-induced EMT. We conclude that papillary thyroid carcinoma tumor cells exhibit increased cancer stem cell-like features during TGF-ß1-induced EMT, that miR-146b-5p has a role in cell proliferation and invasion, and that PRRX1 plays an important role in papillary thyroid carcinoma EMT and disease progression.

Melatonin enhances plant growth and abiotic stress tolerance in soybean plants.

Melatonin is a well-known agent that plays multiple roles in animals. Its possible function in plants is less clear. In the present study, we tested the effect of melatonin (N-acetyl-5-methoxytryptamine) on soybean growth and development. Coating seeds with melatonin significantly promoted soybean growth as judged from leaf size and plant height. This enhancement was also observed in soybean production and their fatty acid content. Melatonin increased pod number and seed number, but not 100-seed weight. Melatonin also improved soybean tolerance to salt and drought stresses. Transcriptome analysis revealed that salt stress inhibited expressions of genes related to binding, oxidoreductase activity/process, and secondary metabolic processes. Melatonin up-regulated expressions of the genes inhibited by salt stress, and hence alleviated the inhibitory effects of salt stress on gene expressions. Further detailed analysis of the affected pathways documents that melatonin probably achieved its promotional roles in soybean through enhancement of genes involved in cell division, photosynthesis, carbohydrate metabolism, fatty acid biosynthesis, and ascorbate metabolism. Our results demonstrate that melatonin has significant potential for improvement of soybean growth and seed production. Further study should uncover more about the molecular mechanisms of melatonin's function in soybeans and other crops.

Viral etiology of community-acquired pneumonia among adolescents and adults with mild or moderate severity and its relation to age and severity.

BACKGROUND: Better knowledge of distribution of respiratory viruses (RVs) in adolescents and adults with community-acquired pneumonia (CAP) is needed. METHODS: To investigate the RVs etiology among adolescents and adults with CAP, according to age and pneumonia severity index (PSI), a multi-center, prospective study was conducted from November 2010 to April 2012. Fifteen RVs were tested by polymerase chain reaction (PCR). Bacteria were detected by urinary antigen, conventional culture and PCR. RESULTS: Mean (SD) age and median (IQR) PSI score of 954 patients enrolled was 45.2 (19.5) years (range 14-94) and 42 (36). RVs were found in 262 patients (27.5%): influenza virus A (IFV A, 9.9%) comprised of pandemic H1N1 (6.7%) and seasonal H3N2 (3.5%), human rhinovirus (4.3%), adenovirus (4.2%), human metapneumovirus (1.8%), parainfluenza virus 1, 3 and 2 (1.7%, 1.5% and 1.2%). Influenza virus B, enterovirus, respiratory syncytial virus, human coronavirus and parainfluenza virus 4 were rarely detected (<1%). Frequency of IFV A was highest among patients aged between 45-64 years (p < 0.001), while adenovirus among patients aged 14-17 years (p < 0.001), no differences was found in other RVs. The proportion of pandemic H1N1 increased with severity of pneumonia evaluated by PSI (P < 0.05). CONCLUSIONS: The proportion of RVs in CAP is higher than previously reported. IFV A pneumonia are usually found in patients older than 45 years, while, adenovirus pneumonia are common in adolescents and young adults. Pandemic H1N1 virus is still recognized by PSI as a high-severity pathogen. The findings contribute baseline data on viral CAP study in China.

Novel approaches in anaplastic thyroid cancer therapy.

Anaplastic thyroid cancer (ATC), accounting for less than 2% of all thyroid cancer, is responsible for the majority of death from all thyroid malignancies and has a median survival of 6 months. The resistance of ATC to conventional thyroid cancer therapies, including radioiodine and thyroid-stimulating hormone suppression, contributes to the very poor prognosis of this malignancy. This review will cover several cellular signaling pathways and mechanisms, including RET/PTC, RAS, BRAF, Notch, p53, and histone deacetylase, which are identified to play roles in the transformation and dedifferentiation process, and therapies that target these pathways. Lastly, novel approaches and agents involving the Notch1 pathway, nuclear factor κB, Trk-fused gene, cancer stem-like cells, mitochondrial mutation, and tumor immune microenvironment are discussed. With a better understanding of the biological process and treatment modality, the hope is to improve ATC outcome in the future.

Hesperetin activates the Notch1 signaling cascade, causes apoptosis, and induces cellular differentiation in anaplastic thyroid cancer.

BACKGROUND: Anaplastic thyroid cancer (ATC) is characterized by very aggressive growth with undifferentiated features. Recently, it has been reported that the Notch1 signaling pathway, which affects thyrocyte proliferation and differentiation, is inactivated in ATC. However, it remains largely unknown whether using Notch1 activating compounds can be an effective therapeutic strategy in ATC. Therefore, in this study, we aimed to evaluate the drug effects of a potential Notch activator hesperetin on ATC cell. METHODS: A unique ATC cell line HTh7 was used to evaluate the drug effects of hesperetin. The Notch1 activating function and cell proliferation were evaluated. The mechanism of growth regulation was investigated by the detection of apoptotic markers. The expression levels of thyrocyte-specific genes were quantified for ATC redifferentiation. RESULTS: Upregulated expression of Notch1 and its downstream effectors hairy and enhancer of split 1 (Hes1) and Hes1 related with YRPW motif was observed in hesperetin-treated ATC cells. The enhanced luciferase signal also confirmed the functional activity of hesperetin-induced Notch1 signaling. Hesperetin led to a time- and dose-dependent decrease in ATC cell proliferation. The cell-growth inhibition was mainly caused by apoptosis as evidenced by increased levels of cleaved poly ADP ribose polymerase and cleaved caspase-3 as well as decreased survivin. Additionally, hesperetin induced the expression levels of thyrocyte-specific genes including thyroid transcription factor 1 (TTF1), TTF2, paired box gene 8, thyroid stimulating hormone receptor, and sodium/iodide symporter. CONCLUSIONS: Hesperetin activates the Notch1 signaling cascade and suppresses ATC cell proliferation mainly via apoptosis. Hesperetin also induces cell redifferentiation of ATC, which could be useful clinically.

The novel histone deacetylase inhibitor thailandepsin A inhibits anaplastic thyroid cancer growth.

BACKGROUND: Anaplastic thyroid cancer (ATC) remains refractory to available surgical and medical interventions. Histone deacetylase (HDAC) inhibitors are an emerging targeted therapy with antiproliferative activity in a variety of thyroid cancer cell lines. Thailandepsin A (TDP-A) is a novel class I HDAC inhibitor whose efficacy remains largely unknown in ATC. Therefore, we aimed to characterize the effect of TDP-A on ATC. METHODS: Human-derived ATC cells were treated with TDP-A. IC50 was determined by a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) rapid colorimetric assay, and cell proliferation was measured by viable cell count. Molecular mechanisms of cell growth inhibition were investigated by Western blot analysis of canonical apoptosis markers, intrinsic and extrinsic apoptosis regulators, and cell cycle regulatory proteins. Cell cycle staging was determined with propidium iodide flow cytometry. RESULTS: TDP-A dose- and time-dependently reduced cell proliferation. Increased cleavage of the apoptosis markers Caspase-9, Caspase-3, and poly adenosine diphosphate ribose polymerase were observed with TDP-A treatment. Levels of the intrinsic apoptosis pathway proteins BAD, Bcl-XL, and BAX remained unchanged. Importantly, the extrinsic apoptosis activator cleaved Caspase-8 increased dose-dependently, and the antiapoptotic proteins Survivin and Bcl-2 decreased. Among the cell cycle regulatory proteins, levels of CDK inhibitors p21/WAF1 and p27/KIP increased. Flow cytometry showed that ATC cells were arrested in G2/M phase with diminished S phase after TDP-A treatment. CONCLUSIONS: TDP-A induces a notable dose- and time-dependent antiproliferative effect on ATC, which is mainly attributed to extrinsic apoptosis with concomitant cell cycle arrest. TDP-A therefore warrants further preclinical and clinical investigations.

Melittin analog p5RHH enhances recombinant adeno-associated virus transduction efficiency.

OBJECTIVE: Melittin and its derivative have been developed to support effective gene delivery systems. Their ability to facilitate endosomal release enhances the delivery of nanoparticle-based gene therapy. Nevertheless, its potential application in the context of viral vectors has not received much attention. Therefore, we would like to optimize the rAAV vector by Melittin analog to improve the transduction efficiency of rAAV in liver cancer cells and explore the mechanism of Melittin analog on rAAV. METHODS: Various melittin-derived peptides were inserted into loop VIII of the capsid protein in recombinant adeno-associated virus vectors. These vectors carrying either gfp or fluc genes were subjected to quantitative polymerase chain reaction assays and transduction assays in human embryonic kidney 293 (HEK293T) cells to investigate the efficiency of vector production and gene delivery. In addition, the ability of a specific p5RHH-rAAV vector to deliver genes was examined through in vitro transduction of different cultured cells and in vivo tail vein administration to C57BL/6 mice. Finally, the intricate details of the vector-mediated transduction mechanisms were explored by using pharmacological inhibitors of every stage of the rAAV2 intracellular life cycle. RESULTS: A total of 76 melittin-related peptides were identified from existing literature. Among them, CMA-3, p5RHH and aAR3 were found to significantly inhibit transduction of rAAV2 vector crude lysate. The p5RHH-rAAV2 vectors efficiently transduced not only rAAV-potent cell lines but also cell lines previously considered resistant to rAAV. Mechanistically, bafilomycin A1, a vacuolar endosome acidification inhibitor, completely inhibited the transgene expression mediated by the p5RHH-rAAV2 vectors. Most importantly, p5RHH-rAAV8 vectors also increased hepatic transduction in vivo in C57BL/6 mice. CONCLUSION: The incorporation of melittin analogs into the rAAV capsids results in a significant improvement in rAAV-mediated transgene expression. While further modifications remain an area of interest, our studies have substantially broadened the pharmacological prospects of melittin in the context of viral vector-mediated gene delivery. Please cite this article as: Meng J, He Y, Yang H, Zhou L, Wang S, Feng X, Al-shargi OY, Yu X, Zhu L, Ling, C. Melittin analog p5RHH enhances recombinant adeno-associated virus transduction efficiency. J Integr Med. 2024; 22(1): 72-82.

Wnt5a inhibits hypoxia-induced pulmonary arterial smooth muscle cell proliferation by downregulation of ß-catenin.

Chronic hypoxia-induced pulmonary arterial hypertension (HPH) is closely associated with profound vascular remodeling, especially pulmonary arterial medial hypertrophy and muscularization due to hyperplasia of pulmonary artery smooth muscle cells (PASMCs). Aberrant Wnt signaling has been associated with lung diseases, but its role in pulmonary hypertension is unclear. This study evaluated the effect of Wnt5a on hypoxia-induced proliferation of human PASMCs and its possible mechanism. The results show that hypoxia (3% O(2), 48 h) induced proliferation of human PASMCs, accompanied with a significant decrease in Wnt5a gene expression, increase in ß-catenin and Cyclin D1 expression, as well as ß-catenin nuclear translocation. Treatment with recombinant mouse Wnt5a significantly inhibited hypoxia-induced proliferation of human PASMCs, upregulation of Cyclin D1 and ß-catenin expression, as well as the nuclear translocation of ß-catenin. These effects were inhibited by Wnt5a antibody. Knocking down ß-catenin or Cyclin D1 gene expression inhibited hypoxia-induced human PASMC proliferation, whereas overexpression of ß-catenin increased hypoxia-induced human PASMC proliferation and counteracted the inhibitory effect of Wnt5a. These results suggest that Wnt5a has an antiproliferative effect on hypoxia-induced human PASMC proliferation by downregulation of ß-catenin and its target gene Cyclin D1. Hypoxia-induced downregulation of Wnt5a may be a way to facilitate hypoxia-induced human PASMC proliferation. The results of this study will help to understand the novel strategies for PH treatment involving Wnt signaling.

Chrysin activates Notch1 signaling and suppresses tumor growth of anaplastic thyroid carcinoma in vitro and in vivo.

BACKGROUND: Anaplastic thyroid cancer (ATC) is a very aggressive thyroid gland malignancy with very poor prognosis. It is suspected that the Notch signaling pathway, which is not active in ATC, may have a tumor suppressor function in this neoplasm. However, it remains unknown whether activation of Notch can yield therapeutic efficacies in ATC. METHODS: The purpose of this study was to evaluate the effect of chrysin, a potential Notch inducer identified via high-throughput screening, on ATC both in vitro and in vivo. RESULTS: Chrysin treatment of ATC cells led to a dose-dependent inhibition of cellular growth. Protein and messenger RNA levels of Notch1 and Hes1 (hairy/enhancer of split 1), a downstream Notch1 effector, were both up-regulated with treatment. Luciferase reporter assays incorporating the C promoter-binding factor 1 (CBF1) binding site also confirmed the functional activity of chrysin-induced Notch1. Oral administration of chrysin suppressed the growth of ATC xenografts by an average of 59% compared with the vehicle control group (P = .002). In addition, calculated median time to tumor progression was 11 days for control mice and 21 days for the chrysin treatment group (P = .008). Analysis of chrysin-treated ATC tumors revealed an increase in the active intracellular domain of Notch1 protein. Activation of Notch1 in vivo was associated with the induction of cleaved Poly ADP ribose polymerase (PARP) protein, indicating that the growth inhibition was due to apoptosis. CONCLUSIONS: The novel Notch1 activator chrysin inhibits tumor growth in ATC both in vitro and in vivo. Chrysin could be a promising therapeutic candidate for ATC, and this justifies further clinical studies.

The use of melittin to enhance transgene expression mediated by recombinant adeno-associated virus serotype 2 vectors both in vitro and in vivo.

OBJECTIVE: Melittin, a cell-penetrating peptide, improves the efficiency of many non-viral gene delivery vectors, yet its application in viral vectors has not been well studied. The non-pathogenic recombinant adeno-associated virus (rAAV) vector is an ideal in vivo gene delivery vector. However, its full potential will only be achieved after improvement of its transduction efficiency. To improve the transduction efficiency of rAAV2 vectors, we attempted to develop a melittin-based rAAV2 vector delivery strategy. METHODS: The melittin peptide was inserted into the rAAV2 capsid either in the loop VIII of all viral proteins (VPs) or at the N terminus of VP2. Various rAAV2-gfp or -fluc vectors were subjected to quantitative real-time polymerase chain reaction and Western blot assays to determine their titers and integrity of capsid proteins, respectively. Alternatively, the vectors based on wild-type capsid were pre-incubated with melittin, followed by transduction of cultured cells or tail vein administration of the mixture to C57BL/6 and BALB/c nude mice. In vivo bioluminescence imaging was performed to evaluate the transgene expression. RESULTS: rAAV2 vectors with melittin peptide inserted in the loop VIII of VPs had low transduction efficiency, probably due to dramatically reduced ability to bind to the target cells. Fusing the melittin peptide at the N-terminus of VP2 produced vectors without the VP2 subunit. Interestingly, among the commonly used rAAV vectors, pre-incubation of rAAV2 and rAAV6 vectors with melittin significantly enhanced their transduction efficiency in HEK293 and Huh7 cells in vitro. Melittin also had the ability to increase the rAAV2-mediated transgene expression in mouse liver in vivo. Mechanistically, melittin did not change the vector-receptor interaction. Moreover, cell counting kit-8 assays of cultured cells and serum transaminase levels indicated melittin had little cytotoxicity. CONCLUSION: Pre-incubation with melittin, but not insertion of melittin into the rAAV2 capsid, significantly enhanced rAAV2-mediated transgene expression. Although further in vivo evaluations are required, this research not only expands the pharmacological potential of melittin, but also provides a new strategy to improve gene therapy mediated by rAAV vectors.

Current treatment of papillary thyroid microcarcinoma.

PTMC is being diagnosed with increasing frequency and generally has an excellent prognosis with less than 0.5% disease-specific mortality. Better prognostic stratification, especially for high-risk patients, helps to optimize surgical care. Older age, extrathyroidal invasion, lymph node involvement, and distant metastases are usually regarded as the most potent risk factors for patients with PTMC. Total or near-total thyroidectomy is advocated as the initial therapy for most primary PTMCs, whereas neck dissection is only recommended with the presence of cervical lymphadenopathy or T4 tumors. ATA suggests that postoperative RAI ablations be administrated to patients with gross extrathyroidal invasion or distant metastases. RAI ablation may also facilitate the use of serum Tg concentrations for postoperative risk assessment.

Should all papillary thyroid microcarcinomas be aggressively treated? An analysis of 18,445 cases.

OBJECTIVE: The purpose of this study was to identify the risk factors that predict papillary thyroid microcarcinoma (PTMC)-related death in a large patient population to determine which patients need aggressive treatment. BACKGROUND: The management of PTMC is controversial and ranges from observation to total thyroidectomy. The lack of consensus is predominantly due to the general excellent overall prognosis, thereby requiring a large cohort to delineate differences in outcome. METHODS: All papillary thyroid cancer patients with tumor size of 1 cm or less in the Surveillance, Epidemiology and End Results Cancer Database from 1988 to 2007 were identified. Outcomes, including overall and disease-specific survival, were compared, and different risk groups were evaluated by multivariate analysis. RESULTS: A total of 18,445 cases of PTMC with surgery were identified. The 10-year and 15-year overall survivals were 94.6% and 90.7%, respectively, while disease-specific survivals were 99.5% and 99.3%. Age greater than 45 years, male sex, African American or minority race, node metastases, extrathyroidal invasion, and distant metastases were stratified to be significant risk factors for overall survival. There were 49 thyroid cancer-related deaths. Forty-five (92%) of the 49 patients had at least 2 risk factors, and 51% of these 49 patients had 3 or more risk factors (vs 5.7% in the rest of the cohort, P < 0.001). CONCLUSIONS: Although PTMC is generally associated with an excellent prognosis, 0.5% patients may die of PTMC. The presence of 2 or more risk factors is strongly associated with cancer-related mortality and can help to identify patients who should be considered for more aggressive management.

Antiproliferative effect of chrysin on anaplastic thyroid cancer.

BACKGROUND: Anaplastic thyroid cancer (ATC) is an undifferentiated, aggressive malignancy, for which there are no effective therapies. Though ATCs only make up less than 2% of all thyroid cancer cases, they represent over half of the thyroid cancer-related deaths. Chrysin, a natural flavonoid, has recently been reported as a potential anti-cancer agent. However, the effect of this compound on ATC cells is not known. Thus, in this study, we evaluated the antiproliferative nature of chrysin in ATC cells. METHODS: HTH7 and KAT18 cells, derived from patients with ATC, were treated with chrysin (25-50 µM) for up to 6 d. Cell proliferation was measured every 2 d using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). Western blot analysis for molecular makers of apoptosis was carried out to investigate the effect and mechanism of Chrysin on ATC. RESULTS: Chrysin inhibited proliferation of HTH7 and KAT18 in a dose- and time-dependent manner. HTH7 and KAT18 cells with Chrysin treatment showed a significant increase in cleaved caspase-3, cleaved PolyADP Ribose Polymerase (PARP), along with a decrease in cyclin D1, Mcl-1, and XIAP. Furthermore, the ratio of Bax to Bcl-2 expression in ATC cells revealed an increase after the treatment. CONCLUSIONS: Chrysin inhibits growth in ATC cells via apoptosis in vitro. Therefore, the natural flavonoid chrysin warrants further clinical investigation as a new potential drug for the treatment for ATC.