Clinicopathologic and molecular correlates to neoadjuvant chemotherapy-induced pathologic response in breast angiosarcoma.

Both primary and secondary breast angiosarcoma (AS) are characterized by multifocal presentation and aggressive behavior. Despite multimodality therapy, local and distant relapse rates remain high. Therefore, neoadjuvant chemotherapy (NACT) is employed to improve the R0 resection rates and survival, but its benefits remain controversial. Herein, we investigate pathologic and molecular correlates to NACT-induced histologic response in a group of 29 breast AS, 4 primary and 25 radiation-associated (RA). The two NACT regimens applied were anthracycline- and non-anthracycline-based. The pathologic response grade was defined as: I: ≤ 50%, II: 51%-90%, III: 91%-99%, and IV: 100%. An additional 45 primary AS and 102 RA-AS treated by surgery alone were included for survival comparison. The genomic landscape was analyzed in a subset of cases and compared to a cohort of AS without NACT on a paired tumor-normal targeted DNA NGS platform. All patients were females, with a median age of 31 years in primary AS and 68 years in RA-AS. All surgical margins were negative in NACT group. The NACT response was evenly divided between poor (Grades I-II; n = 15) and good responders (Grades III-IV; n = 14). Mitotic count >10/mm2 was the only factor inversely associated with pathologic response. By targeted NGS, all 10 post-NACT RA-AS demonstrated MYC amplification, while both primary AS harbored KDR mutations. TMB or other genomic alterations did not correlate with pathologic response. All four patients with Grade IV response remained free of disease. The good responders had a significantly better disease-specific survival (p = 0.04). There was no survival difference with NACT status or the NACT regimens applied. However, NACT patients with MYC-amplified tumors showed better disease-free survival (p = 0.04) compared to MYC-amplified patients without NACT. The overall survival of NACT group correlated with size >10 cm (p = 0.02), pathologic response (p = 0.04), and multifocality (p = 0.01) by univariate, while only size >10 cm (p = 0.03) remained significant by multivariate analysis.

Hinokitiol Inhibits Breast Cancer Cells In Vitro Stemness-Progression and Self-Renewal with Apoptosis and Autophagy Modulation via the CD44/Nanog/SOX2/Oct4 Pathway.

Breast cancer (BC) represents one of the most prevalent malignant threats to women globally. Tumor relapse or metastasis is facilitated by BC stemness progression, contributing to tumorigenicity. Therefore, comprehending the characteristics of stemness progression and the underlying molecular mechanisms is pivotal for BC advancement. Hinokitiol (ß-thujaplicin), a tropolone-related compound abundant in the heartwood of cupressaceous plants, exhibits antimicrobial activity. In our study, we employed three BC cell lines (MDA-MB-231, MCF-7, and T47D) to assess the expression of stemness-, apoptosis-, and autophagy-related proteins. Hinokitiol significantly reduced the viability of cancer cells in a dose-dependent manner. Furthermore, we observed that hinokitiol enhances apoptosis by increasing the levels of cleaved poly-ADP-ribose polymerase (PARP) and phospho-p53. It also induces dysfunction in autophagy through the upregulation of LC3B and p62 protein expression. Additionally, hinokitiol significantly suppressed the number and diameter of cancer cell line spheres by reducing the expression of cluster of differentiation44 (CD44) and key transcription factors. These findings underscore hinokitiol's potential as a therapeutic agent for breast cancer, particularly as a stemness-progression inhibitor. Further research and clinical studies are warranted to explore the full therapeutic potential of hinokitiol in the treatment of breast cancer.

New Histopathologic Risk Model for Early T-stage Oral Squamous Cell Carcinoma: Focusing on a Modified Worst Pattern of Invasion System and a New Tumor Budding Score.

Oral squamous cell carcinoma (OSCC) is treated based on the TNM staging. However, early T-stage OSCC still exhibits substantial nodal metastasis and death rates. Recent literature highlights the independent prognostic value of worst pattern of invasion (WPOI) and tumor budding in OSCC. Nevertheless, WPOI-5 is uncommon in early T-stage OSCC, and the definitions of tumor budding and WPOI-4 overlap. Moreover, WPOI assessment is subjective, and tumor budding evaluation varies across studies. To address these limitations, we aimed to develop a modified WPOI system and a novel tumor budding scoring system that assesses single cells and high-density tumor budding. We also evaluated a new histopathologic risk model for early T-stage OSCC. The study cohort comprised 37 pT1 and 64 pT2 OSCCs. The modified WPOI demonstrated superior interobserver agreement compared with the original system (κ value: 0.98 vs. 0.53). In the multivariate analysis, modified WPOI and tumor budding score were independent prognostic factors for nodal metastasis and disease-free survival, while modified WPOI predicted disease-specific survival. By integrating these factors, our risk model stratified the patients into 3 groups. Notably, the intermediate-risk and high-risk groups exhibited significantly higher rates of nodal metastasis, recurrence, and tumor-related death. Conversely, none in the low-risk group had nodal metastasis or succumbed to the disease. Our model offered simplified scoring and potentially improved prognostic predictions. In conclusion, we've developed a modified WPOI system, a new tumor budding scoring system, and a reliable risk model that classifies early T-stage OSCC patients into distinct risk groups with significant prognostic differences.

Sarcomas With RAD51B Fusions Are Associated With a Heterogeneous Phenotype.

RAD51B-rearranged sarcomas are rare neoplasms that exhibit a heterogeneous morphology. To date, 6 cases have been reported, all involving the uterus, including 4 perivascular epithelioid cell tumors (PEComas) and 2 leiomyosarcomas (LMS). In this study, we describe the morphologic, immunohistochemical, and molecular features of 8 additional sarcomas with RAD51B rearrangement, including the first extrauterine example. All patients were women with a median age of 57 years at presentation. Seven tumors originated in the uterus, and one in the lower extremity soft tissue, with a median tumor size of 12 cm. Histologically, 4 tumors showed predominantly spindle cell morphology with eosinophilic fibrillary cytoplasm, with or without nuclear pleomorphism, whereas 2 tumors exhibited pleomorphic epithelioid cells, featuring clear to eosinophilic, granular cytoplasm. Two neoplasms exhibited undifferentiated cytomorphology, including one with uniform small blue round cells. All tumors showed high-grade cytologic atypia and high mitotic activity (median: 30/10 high-power fields), whereas coagulative necrosis was noted in 6 cases and lymphovascular invasion in 2. By immunohistochemistry, 2 showed myoid and melanocytic markers in keeping with PEComa, whereas 4 cases were only positive for smooth muscle markers consistent with LMS (including 3 myxoid). The remaining 2 cases had a nonspecific immunoprofile. Five cases tested by targeted RNA sequencing (Archer FusionPlex, Illumina TruSight) showed different fusion partners (HMGA2, PDDC1, and CEP170). RAD51B rearrangements were identified by FISH in the remaining 3 cases. Targeted DNA sequencing in 2 cases was negative for TSC gene alterations. Clinical outcome, available in 5 patients (median follow-up, 19 months), revealed 3 local recurrences, 2 lung metastases, and 4 deaths due to disease. Our results expand the spectrum of sarcomas with RAD51B fusions, demonstrating variable clinical presentations, morphologic spectrum, and fusion partners. These tumors have a predilection for a uterine location, with either LMS, PEComa, or undifferentiated phenotypes, and are associated with an aggressive clinical course.

Para-toluenesulfonamide, a novel potent carbonic anhydrase inhibitor, improves hypoxia-induced metastatic breast cancer cell viability and prevents resistance to αPD-1 therapy in triple-negative breast cancer.

Overexpression of the hypoxia-induced transmembrane enzyme carbonic anhydrase IX (CA9) has been associated with poor prognosis and chemoresistance in aggressive breast cancer. This study aimed to investigate the involvement of CA9 in the anti-tumor activity of para-toluenesulfonamide (PTS) and elucidate its mechanism of action against breast cancer both in vitro and in vivo. MCF-7 and MDA-MB-231 breast cancer cells were treated with PTS or subjected to hypoxic conditions using cobalt chloride (CoCl2), with acetazolamide serving as a positive control. Additionally, 4T1 breast cancer cell allograft mice were co-treated with PTS and α-programmed cell death 1 (αPD-1) monoclonal antibody for one month. The results demonstrated that PTS effectively reduced cell viability and reversed migration ability in MCF-7 and MDA-MB-231 cells under CoCl2-induced hypoxia. Furthermore, PTS upregulated the expression of apoptosis-related proteins and downregulated CA9, hypoxia-inducible factor-1α (HIF-1α), and vascular endothelial growth factor (VEGF) proteins, possibly through modulation of p38 MAPK and ERK1/2 phosphorylated proteins. In the animal model, PTS100 inhibited tumor growth and lung metastasis in mammary tumor allograft mice, exhibiting synergistic effects when combined with αPD-1 therapy. Collectively, our findings suggest that PTS inhibits breast cancer growth and metastasis through the p38 MAPK/ERK1/2 pathway. Moreover, PTS may have the potential to prevent the development of resistance to αPD-1 therapy in breast cancer.

Oxidative stress mediates the inhibitory effects of Manzamine A on uterine leiomyoma cell proliferation and extracellular matrix deposition via SOAT inhibition.

Uterine fibroids, the most common benign tumors of the myometrium in women, are characterized by abnormal extracellular matrix deposition and uterine smooth muscle cell neoplasia, with high recurrence rates. Here, we investigated the potential of the marine natural product manzamine A (Manz A), which has potent anti-cancer effects, as a treatment for uterine fibroids. Manz A inhibited leiomyoma cell proliferation in vitro and in vivo by arresting cell cycle progression and inducing caspase-mediated apoptosis. We performed target prediction analysis and identified sterol o-acyltransferases (SOATs) as potential targets of Manz A. Cholesterol esterification and lipid droplet formation were reduced by Manz A, in line with reduced SOAT expression. As a downstream target of SOAT, Manz A also prevented extracellular matrix deposition by inhibiting the ß-catenin/fibronectin/metalloproteinases axis and enhanced autophagy turnover. Excessive free fatty acid accumulation by SOAT inhibition led to reactive oxygen species to impair mitochondrial oxidative phosphorylation and trigger endoplasmic reticulum stress via PERK/eIF2α/CHOP signaling. The inhibitory effect of ManzA on cell proliferation was partially restored by PERK knockdown and eliminated by tauroursodeoxycholic acid, suggesting oxidative stress plays a critical role in the mechanism of action of Manz A. These findings suggest that targeting SOATs by Manz A may be a promising therapeutic approach for uterine fibroids.

The Adipokine Visfatin Modulates Cancer Stem Cell Properties in Triple-Negative Breast Cancer.

Obesity is a cancer progression risk factor; excessive adipocytes increase adipokine secretion. Visfatin, a novel adipokine highly expressed in cancer patients, is related to breast cancer risk. The modulation of nicotinamide adenine dinucleotide (NAD+) metabolism and the induction of a tumorigenic environment plays a vital role in cancer progression. Among cancer cell types, cancer stem-like cells (CSCs) with self-renewal and chemotherapy-resistance abilities could modulate tumor progression and cancer recurrence ability. In this study, we focused on visfatin's modulation effect on stemness-related properties using the high-malignancy breast cancer cell line MDA-MB-231 in in vitro and in vivo studies. Visfatin treatment significantly increased both the sphere number and sphere diameter and increased the protein expression of NANOG homeobox (NANOG), sex-determining region Y-box 2 (SOX2), and octamer-binding transcription factor 4 (OCT4), as well as SIRT1 protein levels. The serum angiogenesis marker VEGF and extracellular nicotinamide phosphoribosyl transferase (NAMPT, visfatin) were induced after visfatin treatment, increasing the stemness and angiogenesis environment, which were significantly reduced by the visfatin inhibitor FK866. Our results demonstrate that the visfatin-activated SIRT-SOX2 axis promotes triple-negative breast cancer stemness and enriches the tumorigenic microenvironment.

ADAD2 functions in spermiogenesis and piRNA biogenesis in mice.

BACKGROUND: Adenosine deaminase domain containing 2 (ADAD2) is a testis-specific protein composed of a double-stranded RNA binding domain and a non-catalytic adenosine deaminase domain. A recent study showed that ADAD2 is indispensable for the male reproduction in mice. However, the detailed functions of ADAD2 remain elusive. OBJECTIVES: This study aimed to investigate the cause of male sterility in Adad2 mutant mice and to understand the molecular functions of ADAD2. MATERIALS AND METHODS: Adad2 homozygous mutant mouse lines, Adad2-/- and Adad2Δ/Δ , were generated by CRISPR/Cas9. Western blotting and immunohistochemistry were used to reveal the expression and subcellular localization of ADAD2. Co-immunoprecipitation tandem mass spectrometry was employed to determine the ADAD2-interacting proteins in mouse testes. RNA-sequencing analyses were carried out to analyze the transcriptome and PIWI-interacting RNA (piRNA) populations in wildtype and Adad2 mutant testes. RESULTS: Adad2-/- and Adad2Δ/Δ mice exhibit male-specific sterility because of abnormal spermiogenesis. ADAD2 interacts with multiple RNA-binding proteins involved in piRNA biogenesis, including MILI, MIWI, RNF17, and YTHDC2. ADAD2 co-localizes and forms novel granules with RNF17 in spermatocytes. Ablation of ADAD2 impairs the formation of RNF17 granules, decreases the number of cluster-derived pachytene piRNAs, and increases expression of ping-pong-derived piRNAs. DISCUSSION AND CONCLUSION: In collaboration with RNF17 and other RNA-binding proteins in spermatocytes, ADAD2 directly or indirectly functions in piRNA biogenesis.

Structure-based virtual screening discovers novel kidney-type glutaminase inhibitors.

Glutaminase (GLS) serves a critical bioenergetic role for malignant tumor growth and has become a valuable therapeutic target for cancer treatment. Herein, we performed a structure-based virtual screening to discover novel GLS inhibitors and provide information for developing new GLS inhibitors. We identified critical pharmacological interactions in the GLS1 binding site by analyzing the known GLS1 inhibitors and selected potential inhibitors based on their docking score and pharmacological interactions. The inhibitory effects of compounds were further confirmed by enzymatic and cell viability assays. We treated colorectal cancer and triple-negative breast cancer cells with the selected candidates and measured the inhibitory efficacy of hit compounds on cell viability. In total, we identified three GLS1 inhibitors. The compounds identified from our structure-based virtual screening methodology exhibited great anticancer potential as a lead targeting glutamine metabolism.

Quantitative Proteome Analysis Reveals Melissa officinalis Extract Targets Mitochondrial Respiration in Colon Cancer Cells.

Melissa officinalis (MO), known as lemon balm, is a popular ingredient blended in herbal tea. In recent decades, the bioactivities of MO have been studied in sub-health and pathological status, highlighting MO possesses multiple pharmacological effects. We previously showed that hot water MO extract exhibited anticancer activity in colorectal cancer (CRC). However, the detailed mechanisms underlying MO-induced cell death remain elusive. To elucidate the anticancer regulation of MO extract in colon cancer, a data-driven analysis by proteomics approaches and bioinformatics analysis was applied. An isobaric tandem mass tags-based quantitative proteome analysis using liquid chromatography-coupled tandem mass spectrometry was performed to acquire proteome-wide expression data. The over-representation analysis and functional class scoring method were implemented to interpret the MO-induced biological regulations. In total, 3465 quantifiable proteoforms were identified from 24,348 peptides, with 67 upregulated and 54 downregulated proteins in the MO-treated group. Mechanistically, MO impeded mitochondrial respiratory electron transport by triggering a reactive oxygen species (ROS)-mediated oxidative stress response. MO hindered the mitochondrial membrane potential by reducing the protein expression in the electron transport chain, specifically the complex I and II, which could be restored by ROS scavenger. The findings comprehensively elucidate how MO hot water extract activates antitumor effects in colorectal cancer (CRC) cells.

Adlay Seed (Coix lacryma-jobi L. var. Ma-yuen Stapf.) Ethanolic Extract Fractions and Subfractions Induce Cell Cycle Arrest and Apoptosis in Human Breast and Cervical Cancer Cell Lines.

The antitumor effects of Coix lacryma-jobi L. var. ma-yuen Stapf. (adlay seed) ethanolic extract have been increasingly shown. This study aimed to investigate the beneficial effects of both the fractions and subfractions of adlay seed ethanolic extract on the human breast (MCF-7) and cervical (HeLa) cancer cell lines, as well as exploring their possible mechanisms of action. The ethanolic extracts were obtained from different parts of adlay seed, including AHE (adlay hull extract), ATE (adlay testa extract), ABE (adlay bran extract) and PAE (polished adlay extract). The results of a 3-(4,5-dimethyl thiazol-2-yl)-2,5-diphenyl- tetrazolium bromide (MTT) assay showed that AHE-Ea and ATE-Ea showed significant growth inhibitory effects in a dose-dependent manner. The results also showed that the AHE-Ea-K, AHE-Ea-L, ATE-Ea-E and ATE-Ea-F subfractions inhibited cell proliferation, induced cell cycle arrest in the G0/G1 phase and decreased CDK4/Cyclin D1 protein expression. Finally, the extract activated caspase-3 activity and PARP protein expression, which induced MCF-7 and HeLa cell apoptosis. We then used liquid chromatography-mass spectrometry (LC/MS) to identify the potential active components., Quercetin showed an anticancer capacity. In conclusion, the AHE-Ea-K, AHE-Ea-L, ATE-Ea-E and ATE-Ea-F subfractions showed antitumor effects through the inhibition of MCF-7 and HeLa cell line viability, as well as inducing apoptosis and cell cycle arrest.

Risk factors for ocular neovascularization after central retinal artery occlusion.

BACKGROUND: To report the incidence and risk factors associated with ocular neovascularization (NV) in patients with central retinal artery occlusion (CRAO). METHODS: This retrospective study included patients diagnosed with acute CRAO in a single tertiary center. Medical charts were reviewed for ocular NV occurrences. We analyzed systemic and ocular conditions on first visit and demographic data. RESULTS: Eighty-seven eyes were eligible for this study. Among these, 13 eyes had ocular NV after CRAO, with an incidence of 15%. The prevalences of hypertension, diabetes mellitus, history of stroke, chronic kidney disease (CKD), and age at first visit were higher among patients with ocular NV than among patients without ocular NV after CRAO. Moreover, most patients with CKD in the ocular NV group had undergone dialysis. A multivariate regression analysis revealed that CKD (hazard ratio [HR]: 9.27, 95% CI, 1.87-46.05, p = 0.006) and glaucoma history (HR: 7.52, 95% CI, 1.14-49.46, p = 0.036) were significant risk factors for developing ocular NV among patients with CRAO. CONCLUSION: CKD and glaucoma history were significant risk factors for developing ocular NV after CRAO, particularly among patients that underwent dialysis.

Spirulina phycocyanin extract and its active components suppress epithelial-mesenchymal transition process in endometrial cancer via targeting TGF-beta1/SMAD4 signaling pathway.

Metastasis is a major challenge in aggressive endometrial cancer treatment accounting for the high recurrence risk and poor prognosis of epithelial-mesenchymal transition (EMT), regulated by the transforming growth factor beta (TGFß) signaling pathway, facilitates tumor metastasis. Spirulina phycocyanin extract (SPE) and its purified products allophycocyanin (APC) and C-phycocyanin (C-PC), derived from Spirulina platensis, can be considered a nutraceutical compound with the ability to inhibit tumor growth and metastasis. Current study aims to investigate the anti-metastatic potential of SPE, and its purified products APC, and C-PC on endometrial cancer both in vitro and in vivo. Firstly, human endometrial cancer cell lines (HEC-1A and Ishikawa) as an in vitro model. Secondly, HEC-1A cells transfected with luminescence gene were implanted into female nude mice as a xenograft model. MTT assay, transwell migration assay, immunoblotting assay, quantitative real-time polymerase chain reaction assay, and IVIS XRMS analysis techniques were used. The in vitro results showed that SPE and its purified products APC and C-PC inhibited cell migration, and altered the expression of EMT-related phenotypes by reversing the TGFß/SMADs signaling pathway. The in vivo results indicated that SPE repressed the metastasis of HEC-1A-LUC cells through modulating EMT-related markers expression. Overall, SPE and its efficient components APC and C-PC reversed the EMT through targeting the TGFß/SMADs signaling pathway, suggesting an effective therapeutic strategy for metastatic endometrial cancer.

Three-Year Outcomes of Intravitreal Aflibercept Injections for Retinal Angiomatous Proliferation According to Disease Stage.

INTRODUCTION: To evaluate the outcomes of intravitreal aflibercept injections for retinal angiomatous proliferation (RAP) according to disease stage. METHODS: This retrospective chart review included 68 eyes of 53 individuals diagnosed as having RAP and 109 neovascular age-related macular degeneration (nAMD) eyes of 109 patients as controls. All patients received intravitreal injections of aflibercept in a real-world setting. The main outcome measures were the changes in the mean of best-corrected visual acuity (BCVA) and central retinal thickness (CRT) as well as the total number of injections received during the 3-year follow-up period. RESULTS: The average BCVA and CRT changes in eyes affected by RAP and the controls at 3 years were non-significant. Both populations received a similar number of injections. After 3 years of treatment, patients with RAP had visual decline despite stable anatomical outcomes. Approximately 50% of the eyes with stage II RAP exhibited significant BCVA decline at the end of the third year. Among those eyes that had deteriorated BCVA, persistently worsening BCVA and thinning CRT were observed from year 2 to year 3. CONCLUSION: Similar to treating nAMD, intensive injections or aggressive treatment strategies are required to treat RAP to achieve optimal visual outcomes in a real-world setting. The response to aflibercept treatment at the second year is associated with the final visual outcome of eyes with stage II RAP lesions.

Screening Tools for Child Abuse Used by Healthcare Providers: A Systematic Review.

BACKGROUND: The early detection of child abuse is essential for children at risk. Healthcare professionals working at hospitals and in the community are often the first to encounter suspected cases of child abuse. Therefore, an accurate identification of child abuse is critical for intervention. However, there is no consensus on the best method to screen for child abuse. PURPOSE: This systematic review was designed to evaluate the relevant psychometric properties and critically appraise the methodological quality of child abuse screening tools used by healthcare providers with children less than 18 years old. METHODS: We searched the Cochrane Library, MEDLINE, Embase, CINAHL, Education Resources Information Center, PubMed, Airiti Library, and OpenGray databases for studies on screening tools used to identify abuse in children published through October 2019 in English or Chinese. Information regarding populations, assessment methods, and accuracy parameters were extracted. Study quality was assessed using the COnsensus-based Standards for the selection of health Measurement INstruments checklist and Grading of Recommendation, Assessment, Development, and Evaluation criteria. RESULTS: Nine hundred thirty-nine abstracts and 23 full-text articles were reviewed for eligibility, and 15 screening tools for child abuse used by healthcare providers were identified. Screening tools often assess the presence of more than one form of abuse, but no single tool covered all forms. Of these, 10 tools screened for a single, discrete type of abuse, including nine physical abuse screening tools (three abusive head trauma tools) and one sexual abuse tool. Eighty percent (n = 12) of the screening tools had a moderate-to-high quality of evidence based on the Grading of Recommendation, Assessment, Development, and Evaluation criteria. However, none of these screening tools achieved an adequate level of evidence based on the COnsensus-based Standards for the selection of health Measurement INstruments checklist. CONCLUSIONS/IMPLICATIONS FOR PRACTICE: In this systematic literature review, 15 assessment tools of child abuse used by healthcare providers were identified, of which nine screened for physical abuse. Screening tools must be valid, succinct, user-friendly, and amenable for use with children at every point of care in the healthcare system. Because of the paucity of informative and practical studies in the literature, findings related to the quality of child abuse screening tools were inconclusive. Therefore, future research should focus on the use of screening tools in the healthcare system to identify effective screening interventions that may help healthcare providers identify child abuse cases as early as possible.

Probiotic supplementation attenuates age-related sarcopenia via the gut-muscle axis in SAMP8 mice.

BACKGROUND: Age-related muscle dysfunctions are common disorders resulting in poor quality of life in the elderly. Probiotic supplementation is a potential strategy for preventing age-related sarcopenia as evidence suggests that probiotics can enhance muscle function via the gut-muscle axis. However, the effects and mechanisms of probiotics in age-related sarcopenia are currently unknown. In this study, we examined the effects of Lactobacillus casei Shirota (LcS), a probiotic previously reported to improve muscle function in young adult mice. METHODS: We administered LcS (1 × 108 or 1 × 109  CFU/mouse/day) by oral gavage to senescence-accelerated mouse prone-8 mice for 12 weeks (16- to 28-week-old). Sixteen-week-old and 28-week-old SMAP8 mice were included as non-aged and aged controls, respectively. Muscle condition was evaluated using dual-energy X-ray absorptiometry for muscle mass, holding impulse and grip strength tests for muscle strength, and oxygen consumption rate, gene expressions of mitochondrial biogenesis, and mitochondrial number assays for mitochondria function. Inflammatory cytokines were determined using enzyme-linked immunosorbent assay. Gas chromatography-mass spectrometry was utilized to measure the short-chain fatty acid levels. The gut microbiota was analysed based on the data of 16S rRNA gene sequencing of mouse stool. RESULTS: The LcS supplementation reduced age-related declines in muscle mass (>94.6%, P < 0.04), strength (>66% in holding impulse and >96.3% in grip strength, P < 0.05), and mitochondrial function (P < 0.05). The concentration of short-chain fatty acids (acetic, isobutyric, butyric, penic, and hexanoic acid) was recovered by LcS (>65.9% in the mice given high dose of LcS, P < 0.05) in the aged mice, and LcS attenuated age-related increases in inflammation (P < 0.05) and reactive oxygen species (>89.4%, P < 0.001). The high dose of LcS supplementation was also associated with distinct microbiota composition as indicated by the separation of groups in the beta-diversity analysis (P = 0.027). LcS supplementation altered predicted bacterial functions based on the gut microbiota. Apoptosis (P = 0.026), p53 signalling (P = 0.017), and non-homologous end-joining (P = 0.031) were significantly reduced, whereas DNA repair and recombination proteins (P = 0.043), RNA polymerase (P = 0.008), and aminoacyl-tRNA biosynthesis (P = 0.003) were increased. Finally, the genera enriched by high-dose LcS [linear discriminant analysis (LDA) score > 2.0] were positively correlated with healthy muscle and physiological condition (P < 0.05), while the genera enriched in aged control mice (LDA score > 2.0) were negatively associated with healthy muscle and physiological condition (P < 0.05). CONCLUSIONS: Lactobacillus casei Shirota represents an active modulator that regulates the onset and progression of age-related muscle impairment potentially via the gut-muscle axis.

Simultaneous determination of areca nut- and tobacco-specific alkaloids in saliva by LC-MS/MS: Distribution and transformation of alkaloids in oral cavity.

Areca nut and tobacco are frequently used in combination. Cigarette smoking and betel quid (BQ) chewing habits impose greater oral cancer risk than either habit alone. Saliva is a better noninvasive diagnostic material as it is in direct contact with oral mucosa and cancerous lesions. This study describes the application of isotope-dilution LC-MS/MS for simultaneous quantitation of five areca nut-specific alkaloids (ASAs) and three tobacco-specific alkaloids (TSAs) in human saliva. With this method, we demonstrate that the distribution of ASAs vary significantly in smokers who chew BQ habitually, due to the hydrolysis of ASAs and metabolic activity in the oral cavity. The alkaline condition formed due to slaked lime in BQ, plays an important role in the distribution of ASAs and TSAs, by catalyzing the hydrolysis of ester forms of ASAs to their respective carboxylic acid forms besides facilitating the TSA (i.e., nicotine) absorption in the oral cavity. Moreover, our results reveal that oral mucosa rather than saliva contributes to the metabolism of ASAs at oral cavity. Less than 2.1% of ASAs were metabolized by saliva, as determined by in vitro test. Our findings may provide a better insight into the pathobiology of oral carcinogenesis due to BQ chewing.

The Role of Cell Proliferation and Extracellular Matrix Accumulation Induced by Food Additive Butylated Hydroxytoluene in Uterine Leiomyoma.

Leiomyoma is the most common benign uterine tumor in reproductive-age women. Increasing numbers of studies are focusing on the effects of environmental exposure on the incidence and progression of tumors. One major step taken in the food industry is the addition of food preservatives to maintain freshness. Butylated hydroxytoluene (BHT) is a synthetic phenolic antioxidant, which is widely used as an additive to develop fat-soluble characteristics, as well as in cosmetics and rubber. Previous studies also highlighted that BHT may be related to increased fibrosis capacity and carcinogenic effects. In this study, we explored the effects of the commonly used food additive BHT on leiomyoma progression, and the related mechanism. The exposure of the ELT-3 leiomyoma cell line to BHT for 48 h increased the proliferative effect. Since leiomyoma progression is related to increases in extracellular matrix (ECM) accumulation and matrix metalloproteinase (MMP), BHT could effectively increase ECM-related protein expression, as well as MMP-2 and MMP-9 protein expression. This increase in ECM, in response to BHT, may be linked to the activation of the phosphoinositide 3-kinase (PI3K)/Akt and mitogen-activated protein kinase (MAPK) signaling pathway. Through PI3K inhibition, BHT's effect on leiomyoma progression could be partially modulated. These results suggest the harmful effect of BHT exposure on leiomyoma progression may relate to PI3K modulation. However, an in vivo study is necessary to confirm these findings.

Predicting Agents That Can Overcome 5-FU Resistance in Colorectal Cancers via Pharmacogenomic Analysis.

5-Fluorouracil (5-FU) is one of several chemotherapeutic agents in clinical use as a standard of care to treat colorectal cancers (CRCs). As an antimetabolite, 5-FU inhibits thymidylate synthase to disrupt the synthesis and repair of DNA and RNA. However, only a small proportion of patients benefit from 5-FU treatment due to the development of drug resistance. This study applied pharmacogenomic analysis using two public resources, the Genomics of Drug Sensitivity in Cancer (GDSC) and the Connectivity Map, to predict agents overcoming 5-FU resistance in CRC cells based on their genetic background or gene expression profile. Based on the genetic status of adenomatous polyposis coli (APC), the most frequent mutated gene found in CRC, we found that combining a MEK inhibitor with 5-FU exhibited synergism effects on CRC cells with APC truncations. While considering the gene expression in 5-FU resistant cells, we demonstrated that targeting ROCK is a potential avenue to restore 5-FU response to resistant cells with wild-type APC background. Our results reveal MEK signaling plays a pivotal role in loss-of-function, APC-mediated 5-FU resistance, and ROCK activation serves as a signature in APC-independent 5-FU resistance. Through the use of these available database resources, we highlight possible approaches to predict potential drugs for combinatorial therapy for patients developing resistance to 5-FU treatment.

Hinokitiol Exhibits Antitumor Properties through Induction of ROS-Mediated Apoptosis and p53-Driven Cell-Cycle Arrest in Endometrial Cancer Cell Lines (Ishikawa, HEC-1A, KLE).

Hinokitiol is a natural tropolone derivative that is present in the heartwood of cupressaceous plants, and has been extensively investigated for its anti-inflammatory, antioxidant, and antitumor properties in the context of various diseases. To date, the effects of hinokitiol on endometrial cancer (EC) has not been explored. The purpose of our study was to investigate the anti-proliferative effects of hinokitiol on EC cells. Cell viability was determined with an MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay, and the quantification of apoptosis and reactive oxygen species (ROSs) was performed by using flow cytometry, while protein expression was measured with the Western blotting technique. Hinokitiol significantly suppressed cell proliferation through the inhibition of the expression of cell-cycle mediators, such as cyclin D1 and cyclin-dependent kinase 4 (CDK4), as well as the induction of the tumor suppressor protein p53. In addition, hinokitiol increased the number of apoptotic cells and increased the protein expression of cleaved-poly-ADP-ribose polymerase (PARP) and active cleaved-caspase-3, as well as the ratio of Bcl-2-associated X protein (Bax) to B-cell lymphoma 2 (Bcl-2). Interestingly, except for KLE cells, hinokitiol induced autophagy by promoting the accumulation of the microtubule-associated protein light chain 3B (LC3B) and reducing the sequestosome-1 (p62/SQSTM1) protein level. Furthermore, hinokitiol triggered ROS production and upregulated the phosphorylation of extracellular-signal-regulated kinase (p-ERK1/2) in EC cells. These results demonstrate that hinokitiol has potential anti-proliferative and pro-apoptotic benefits in the treatment of endometrial cancer cell lines (Ishikawa, HEC-1A, and KLE).