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Cell therapies such as genetically modified T cells have emerged as a promising and viable treatment for hematologic cancers and are being aggressively pursued for a wide range of diseases and conditions that were previously difficult to treat or had no cure. The process development requires genetic modifications to T cells to express a receptor (engineered T cell receptor (eTCR)) of specific binding qualities to the desired target. Protein reagents utilized during the cell therapy manufacturing process, to facilitate these genetic modifications, are often present as process-related impurities at residual levels in the final drug product and can represent a potential immunogenicity risk upon infusion. This manuscript presents a framework for the qualification of an assay for assessing the immunogenicity risk of AA6 and Cas9 residuals. The same framework applies for other residuals; however, AAV6 and Cas9 were selected as they were residuals from the manufacturing of an engineered T cell receptor cellular product in development. The manuscript: 1) elucidates theoretical risks, 2) summarizes analytical data collected during process development, 3) describes the qualification of an in vitro human PBMC cytokine release assay to assess immunogenicity risk from cellular product associated process residuals; 4) identifies a multiplexed inflammatory innate and adaptive cytokine panel with pre-defined criteria using relevant positive controls; and 5) discusses qualification challenges and potential solutions for establishing meaningful thresholds. The assessment is not only relevant to establishing safe exposure levels of these residuals but also in guiding risk assessment and CMC strategy during the conduct of clinical trials.
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Receptores de Antígenos de Linfocitos T , Linfocitos T , Humanos , Medición de Riesgo/métodos , Receptores de Antígenos de Linfocitos T/inmunología , Linfocitos T/inmunología , Citocinas/metabolismo , Leucocitos Mononucleares/inmunología , Leucocitos Mononucleares/metabolismo , Leucocitos Mononucleares/efectos de los fármacos , Contaminación de Medicamentos/prevención & controlRESUMEN
OBJECTIVE: To analyse the aetiology and pathogenesis of Gorlin-Goltz syndrome (GS; also known as nevoid basal cell carcinoma syndrome [NBCCS] or basal cell nevus syndrome [BCNS]) in a Chinese family. METHODS: Whole-exome sequencing (WES) was performed on genomic DNA samples from the subjects in a family, followed by the investigation of pathogenesis via bioinformatic approaches and conformational analysis. RESULTS: A novel heterozygous non-frameshift deletion patched 1 (PTCH1) [NM_000264: c.3512_3526del (p.1171_1176del)] was identified by WES and further validated by Sanger sequencing. Bioinformatic and conformational analysis showed that the mutation caused altered PTCH1 protein structure, which may be related to functional abnormalities. CONCLUSION: This study expands the mutation spectrum of PTCH1 in GS and facilitates the early diagnosis and screening of GS. PTCH1 [c.3512_3526del (p.1171_1176del)] may cause structural abnormalities and functional disabilities, leading to GS in families.
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Síndrome del Nevo Basocelular , Humanos , Síndrome del Nevo Basocelular/genética , Causalidad , Biología Computacional , Mutación , Pueblos del Este de AsiaRESUMEN
Collagen membrane with outstanding biocompatibility exhibits immense potential in the field of corneal repair and reconstruction, but the poor mechanical properties limit its clinical application. Polycaprolactone (PCL) is a biodegradable polymer widely explored for application in corneal reconstruction due to its excellent mechanical properties, biocompatibility, easy processability, and flexibility. In this study, a PCL/collagen composite membrane with reinforced mechanical properties is developed. The membrane has a strong composite structure with collagen by utilizing a porous and hydrophilic PCL scaffold, maintaining its integrity even after immersion. The suture retention and mechanical tests demonstrate that compared with the pure collagen membrane, the prepared membrane has a greater tensile strength and twice the modulus of elasticity. Further, the suture retention strength is improved by almost two times. In addition, the membrane remains fully intact on the implant bed in an in vitro corneal defect model. Moreover, the membrane can be tightly sutured to a rabbit corneal defect, progressively achieve epithelialization, and remain unchanged during observation. Overall, the PCL/collagen composite membrane is a promising candidate as a suturable corneal restoration material in clinical keratoplasty.
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Colágeno , Córnea , Poliésteres , Animales , Conejos , Colágeno/química , Poliésteres/química , Porosidad , Resistencia a la Tracción , Membranas Artificiales , Interacciones Hidrofóbicas e Hidrofílicas , Ensayo de Materiales , Materiales Biocompatibles/química , Materiales Biocompatibles/farmacologíaRESUMEN
OBJECTIVE: We aimed to develop an electroactive antibacterial material for the treatment of skin wound diseases. METHOD: To this aim, we modified chitosan (CS), a biocompatible polymer, by coupling it with graphene (rGO) and an antimicrobial polypeptide DOPA-PonG1. The material's effect on skin injury healing was studied in combination with external electrical stimulation (EEM). The structure, surface composition, and hydrophilicity of the modified CS materials were evaluated using scanning electron microscopy (SEM), Fourier-transform infrared spectroscopy (FTIR), and contact angle measurements. We studied NIH3T3 cells cultured with modified materials and subjected to EEM to assess viability, adhesion, and tissue repair-related gene expression. RESULTS: SEM data demonstrated that rGO was distributed uniformly on the surface of the CS material, increasing surface roughness, and antimicrobial peptides had minimal impact on surface morphology. FTIR confirmed the uniform distribution of rGO and antibacterial peptides on the material surface. Both rGO and DOPA-PonG1 enhanced the hydrophilicity of CS materials, with rGO also improving tensile strength. The dual modification of CS with rGO and DOPA-PonG1 synergistically increased antibacterial efficacy. Cellular events and gene expression relevant to tissue repair process were enhanced by these modifications. Furthermore, EEM accelerated epidermal regeneration more than the material alone. In a rat skin wound model, DOPA-PonG1@CS/rGO dressing combined with electrical stimulation exhibited accelerated healing of skin defect. CONCLUSION: Overall, our results demonstrate that CS materials modified with rGO and DOPA-PonG1 have increased hydrophilicity, antibacterial characteristics, and tissue regeneration capacities. This modified material in conjunction with EEM hold promise for the clinical management for dermal wounds.
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Antibacterianos , Quitosano , Ratones , Ratas , Animales , Células 3T3 NIH , Antibacterianos/farmacología , Antibacterianos/uso terapéutico , Quitosano/farmacología , Quitosano/química , Estimulación Eléctrica , Vendajes , DihidroxifenilalaninaRESUMEN
Allium mongolicum Regel (A. mongolicum) is a healthy edible plant but highly perishable with a short shelf life of 1-2 d. Modified atmosphere packaging (MAP) could inhibit the postharvest senescence and decay of the vegetables. Thus, the aim of this study was to apply MAP with different gas permeabilities to the storage of A. mongolicum and evaluate its effects on maintaining microbial communities and the postharvest quality of A. mongolicum. The results showed that polypropylene/poly(butylene adipate-co-terephthalate) (PP/PBAT, abbreviated as PAT) MAP was suitable for the storage of A. mongolicum by establishing an optimal atmosphere of 0.5-0.6% O2 and 6.2-7.1% CO2 in the bag. It could delay the postharvest senescence of A. mongolicum and maintain its quality by slowing down its respiration rate and weight loss, reducing cell membrane permeability and lipid peroxidation, maintaining the cell wall, and reducing infection and the growth of microorganisms. However, A. mongolicum in HPT was more perishable than that in PAT during storage. Pseudomonas was found to be the main spoilage bacteria, and they could also be effectively inhibited by PAT-MAP. The next-generation sequencing results also showed the growth of Escherichia-Shigella, Clostridium sensu stricto 1, Streptococcus, Aureobasidium, Didymella, and Fusarium, responsible for A. mongolicum decay or human disease, was well inhibited by PAT-MAP. The results suggested that PAT-MAP could be used to maintain microbial diversity and the postharvest quality of A. mongolicum under cold storage conditions. It provided a feasible solution for the preservation, food quality, and safety control of A. mongolicum.
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BACKGROUND: To develop a radiomics model based on chest computed tomography (CT) for the prediction of a pathological complete response (pCR) after neoadjuvant or conversion chemoimmunotherapy (CIT) in patients with non-small cell lung cancer (NSCLC). METHODS: Patients with stage IB-III NSCLC who received neoadjuvant or conversion CIT between September 2019 and July 2021 at Hunan Cancer Hospital, Xiangya Hospital, and Union Hospital were retrospectively collected. The least absolute shrinkage and selection operator (LASSO) were used to screen features. Then, model 1 (five radiomics features before CIT), model 2 (four radiomics features after CIT and before surgery) and model 3 were constructed for the prediction of pCR. Model 3 included all nine features of model 1 and 2 and was later named the neoadjuvant chemoimmunotherapy-related pathological response prediction model (NACIP). RESULTS: This study included 110 patients: 77 in the training set and 33 in the validation set. Thirty-nine (35.5%) patients achieved a pCR. Model 1 showed area under the curve (AUC) = 0.65, 64% accuracy, 71% specificity, and 50% sensitivity, while model 2 displayed AUC = 0.81, 73% accuracy, 62% specificity, and 92% sensitivity. In comparison, NACIP yielded a good predictive value, with an AUC of 0.85, 81% accuracy, 81% specificity, and 83% sensitivity in the validation set. CONCLUSION: NACIP may be a potential model for the early prediction of pCR in patients with NSCLC treated with neoadjuvant/conversion CIT.
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Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Humanos , Carcinoma de Pulmón de Células no Pequeñas/diagnóstico por imagen , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Neoplasias Pulmonares/diagnóstico por imagen , Neoplasias Pulmonares/tratamiento farmacológico , Terapia Neoadyuvante , Estudios Retrospectivos , Área Bajo la CurvaRESUMEN
Knowledge of the ionome of plant organs helps us understand a plant's nutritional status. However, the ionome of Macadamia (Proteaceae), which is an important nut-producing tree, remains unknown. We aimed to characterise the allocation of biomass and nutrient-partitioning patterns in three macadamia genotypes. We excavated 15 productive trees (three cultivars at 21years of age; two cultivars at 16years of age) in an orchard. Biomass, nutrient concentrations, and contents of roots, stems, branches, and leaves were analysed. Dry weight of roots, stems, branches and leaves accounted for 14-20%, 19-30%, 36-52%, and 12-18% of total plant weight, respectively. No significant difference was found in the total biomass among the cultivars at the same age. Compared with most crop plants, macadamia had low phosphorus (P) concentrations in all organs (<1gkg-1 ), and low leaf zinc (Zn) concentration (8mgkg-1 ). In contrast, macadamia accumulated large amounts of manganese (Mn), with a 20-fold higher leaf Mn concentration than what is considered sufficient for crop plants. Leaves exhibited the highest nutrient concentrations, except for iron and Zn, which exhibited the highest concentrations in roots. The organ-specific ionomics of Macadamia is characterised by low P and high Mn concentrations, associated with adaptation to P-impoverished habitats.
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Macadamia , Proteaceae , Manganeso , Biomasa , Plantas , Árboles , FósforoRESUMEN
BACKGROUND: The present study investigated the regulatory effects of N6-methyladenosine (m6A) methyltransferase like-3 (METTL3) in diabetes-induced testicular damage. METHODS: In vivo diabetic mice and high glucose (HG) treated GC-1 spg cells were established. The mRNA and protein expressions were determined by real-time quantitative polymerase chain reaction, Western blot, immunofluorescence and immunohistochemistry staining. Levels of testosterone, blood glucose, cell viability, and apoptosis were detected by enzyme-linked immunosorbent assay, MTT, and flow cytometry, respectively. Molecular interactions were verified by RNA immunoprecipitation and RNA pull-down assay. Histopathological staining was performed to evaluate testicular injury. RESULTS: METTL3 and long non-coding RNA taurine up-regulated 1 (lncRNA TUG1) were downregulated in testicular tissues of diabetic mice and HG-treated GC-1 spg cells. METTL3 overexpression could reduce the blood glucose level, oxidative stress and testicular damage but enhance testosterone secretion in diabetic mouse model and HG-stimulated GC-1 spg cells. Mechanically, METTL3-mediated m6A methylation enhanced the stability of TUG1, then stabilizing the clusterin mRNA via recruiting serine and arginine rich splicing factor 1. Moreover, inhibition of TUG1/clusterin signaling markedly reversed the protective impacts of METTL3 overexpression on HG-stimulated GC-1 spg cells. CONCLUSION: This study demonstrated that METTL3 ameliorated diabetes-induced testicular damage by upregulating the TUG1/clusterin signaling. These data further elucidate the potential regulatory mechanisms of m6A modification on diabetes-induced testicular injury.
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Diabetes Mellitus Experimental , Metiltransferasas , Animales , Ratones , Glucemia , Clusterina , Diabetes Mellitus Experimental/complicaciones , Metiltransferasas/genética , Metiltransferasas/metabolismo , ARN , ARN Mensajero/genética , ARN Mensajero/metabolismo , TestosteronaRESUMEN
The study was performed to ascertain the mechanism of sodium butyrate (NaB) mediating the proliferative and invasive properties of oral squamous cell carcinoma (OSCC) cells. The cell proliferative, migrating, and invasive potentials were detected by CCK-8, colony formation, EdU, and Transwell assays. The expression of proliferation- and invasion-related proteins, HDAC1, and HSPB7 in OSCC cells were evaluated by western blot. Immunofluorescence was also performed to evaluate the HDAC1 expression. The enrichment of histone deacetylase HDAC1 in the promoter region of HSPB7 was assessed by the ChIP assay. In vivo growth of OSCC cells was measured by tumorigenesis in nude mice (n=18). The t-test was employed for comparisons of data between the two groups. One-way ANOVA was utilized for comparisons of data among multiple groups, and repeated-measures ANOVA for comparisons of data at different time points among groups, followed by Bonferroni post-hoc test. The data showed that HDAC1 expression was highly upregulated in OSCC cells compared to human normal oral keratinocytes (HNOKs) (p<0.0001), and NaB diminished the HDAC1 expression in OSCC cells. NaB restricted OSCC cell proliferative, migrating, and invasive capabilities by downregulating HDAC1. HSPB7 expression was downregulated in OSCC cells versus HNOKs (p<0.0001). HDAC1 inversely orchestrated the HSPB7 expression in OSCC cells through histone deacetylation modification, and NaB augmented the HSPB7 expression by inhibiting HDAC1. Moreover, NaB inhibited OSCC cell growth in vivo by elevating HSPB7 levels through the HDAC1 repression. In conclusion, NaB restrained cell proliferation and invasion in OSCC cells via HSPB7 upregulation by decreasing the HDAC1 expression.
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Carcinoma de Células Escamosas , Neoplasias de Cabeza y Cuello , MicroARNs , Neoplasias de la Boca , Animales , Ácido Butírico/farmacología , Carcinoma de Células Escamosas/genética , Línea Celular Tumoral , Movimiento Celular , Proliferación Celular , Regulación Neoplásica de la Expresión Génica , Proteínas de Choque Térmico HSP27/metabolismo , Histona Desacetilasa 1 , Histona Desacetilasas/metabolismo , Histonas , Humanos , Ratones , Ratones Desnudos , Neoplasias de la Boca/tratamiento farmacológico , Neoplasias de la Boca/genética , Sincalida/metabolismo , Carcinoma de Células Escamosas de Cabeza y CuelloRESUMEN
INTRODUCTION: Periodontal disease and osteoporosis are common chronic diseases, especially for the postmenopausal women. Several original studies explore the association, but there still controversial. Therefore, we will conduct this systematic review and meta-analysis to assess the association between periodontal disease and osteoporosis in postmenopausal women. METHODS AND ANALYSIS: This study adheres to the Preferred Reporting Items for Systematic Reviews and Meta-analyses for Protocols. We will systematically search Medline/PubMed, Embase, Cochrane Central Register of Controlled Trials, Web of Science and Scopus from inception to August 2021 to collect all relevant publications, with no restrictions on publication date or languages. Study selection, data extraction and risk of bias assessment will be conducted independently by two trained reviewers independently. The Cochrane's tool for assessing risk of bias, Newcastle-Ottawa Scale and Agency for Healthcare Research and Quality will be used for the risk of bias assessment. OR, HR and risk ratio with 95% CI were considered as the effect size for dichotomous outcomes, weighted mean diï¬erence with 95% CI were calculated as the effect size for continuous outcomes. Random-effects models will be used. Heterogeneity between studies will be assessed via the forest plot and I². Publication bias will detected by funnel plots, Begg's test and Egger's test. The subgroup analyses and sensitivity ananlyses will also be used to explore and interpret the heterogeneity. ETHICS AND DISSEMINATION: This study does not require ethical approval. We will disseminate our findings by publishing results in a peer-reviewed journal. PROSPERO REGISTRATION NUMBER: CRD42021225746.
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Osteoporosis , Enfermedades Periodontales , Femenino , Humanos , Metaanálisis como Asunto , Enfermedades Periodontales/complicaciones , Enfermedades Periodontales/epidemiología , Posmenopausia , Proyectos de Investigación , Revisiones Sistemáticas como AsuntoRESUMEN
To estimate whether adjuvant radiotherapy is necessary for patients with stage IA1-IIA1 cervical cancer after laparoscopic hysterectomy, 221 patients were retrospectively analyzed. Sixty-two of them were treated with laparoscopic hysterectomy and adjuvant radiotherapy (group A), 115 underwent open surgery (group B) and 44 received laparoscopic hysterectomy alone (group C). Results showed that the 3-year local recurrence-free survival (LRFS) rates of group A, B and C were 98.4%, 97.4% and 86.4%, respectively. The LRFS rates of group A and B surpassed C (A vs. B, p=0.634; A vs. C, p=0.011; B vs. C, p=0.006). The inter-group differences of 3-year overall survival (OS) and distant metastasis free survival (DMFS) were not statistically significant. In subgroup analysis of stage IB disease, the 3-year LRFS rates of group A, B and C were 100%, 98.8% and 83.1%, the 3-year OS rates of group A, B and C were 100%, 98.9% and 91.5%, respectively. The 3-year LRFS and OS rates of group A and B were significantly superior to group C (p<0.05). Our findings suggest that adjuvant radiotherapy can reduce the risk of recurrence for women with early-stage cervical cancer after laparoscopic hysterectomy and bring survival benefits for patients with stage IB disease.
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The coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is spreading at an alarming rate, and it has created an unprecedented health emergency threatening tens of millions of people worldwide. Previous studies have indicated that SARS-CoV-2 ribonucleic acid could be detected in the feces of patients even after smear-negative respiratory samples. However, demonstration of confirmed fecal-oral transmission has been difficult. Clinical studies have shown an incidence rate of gastrointestinal (GI) symptoms ranging from 2% to 79.1% in patients with COVID-19. They may precede or accompany respiratory symptoms. The most common GI symptoms included nausea, diarrhea, and abdominal pain. In addition, some patients also had liver injury, pancreatic damage, and even acute mesenteric ischemia/thrombosis. Although the incidence rates reported in different centers were quite different, the digestive system was the clinical component of the COVID-19 section. Studies have shown that angiotensin-converting enzyme 2, the receptor of SARS-CoV-2, was not only expressed in the lungs, but also in the upper esophagus, small intestine, liver, and colon. The possible mechanism of GI symptoms in COVID-19 patients may include direct viral invasion into target cells, dysregulation of angiotensin-converting enzyme 2, immune-mediated tissue injury, and gut dysbiosis caused by microbiota. Additionally, numerous experiences, guidelines, recommendations, and position statements were published or released by different organizations and societies worldwide to optimize the management practice of outpatients, inpatients, and endoscopy in the era of COVID-19. In this review, based on our previous work and relevant literature, we mainly discuss potential fecal-oral transmission, GI manifestations, abdominal imaging findings, relevant pathophysiological mechanisms, and infection control and prevention measures in the time of COVID-19.
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A pair of stereoisomers of new 4,5-dihydroxypiperine was isolated from P. retrofractum and showed profound activity on AlCl3-induced dementia. In order to determine their absolute configurations and biological activities, all four possible stereoisomers of 4,5-dihydroxypiperine were synthesized from piperidine by Sharpless asymmetric dihydroxylation and Mitsunobu reaction. Their absolute configurations were established as (4R,5R) (1), (4S,5S) (2), (4S,5R) (3) and (4R,5S) (4) by NMR, optical rotation and CD spectra. It is note that only compound 4 improved behavioral disorder in AlCl3-induced dementia. Accordingly, the pair of stereoisomers isolated from P. retrofractum was determined to be (4S,5S) and (4R,5S)-isomers (2 and 4). The ratio of the epimers was present as 1:0.7 (4:2).
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Alcaloides/farmacología , Benzodioxoles/farmacología , Demencia/tratamiento farmacológico , Piperaceae/química , Piperidinas/farmacología , Alcamidas Poliinsaturadas/farmacología , Alcaloides/química , Alcaloides/aislamiento & purificación , Cloruro de Aluminio , Animales , Terapia Conductista , Benzodioxoles/química , Benzodioxoles/aislamiento & purificación , Demencia/inducido químicamente , Relación Dosis-Respuesta a Droga , Estructura Molecular , Piperidinas/química , Piperidinas/aislamiento & purificación , Alcamidas Poliinsaturadas/química , Alcamidas Poliinsaturadas/aislamiento & purificación , Relación Estructura-Actividad , Pez CebraRESUMEN
BACKGROUND INFORMATION: Diabetes-induced testicular dysfunction is characterised by abnormal apoptosis of spermatogenic cells, but the underlying mechanism is poorly understood. This study aimed to investigate the roles of clusterin (CLU) in testicular damage associated with diabetes pathogenesis, as well as the molecular mechanism. A rat diabetes model was established using streptozocin, and the mouse spermatogenic cell line GC-1 spg was treated with high glucose as a cellular model. CLU was overexpressed in GC-1 spg cells, followed by detection of serum testosterone, cell proliferation, cell apoptosis and autophagy. RESULTS: CLU expression was significantly reduced and LC3 expression was elevated in testis tissues in the rat diabetes model and high glucose-treated GC-1 spg cells. High glucose led to suppressed viability, enhanced apoptosis, reduced Bcl-2 expression, elevated Bax expression and cleavage of Caspase-3/-9 in GC-1 spg cells, and these effects were abrogated by CLU overexpression. Additionally, CLU overexpression repressed LC3 and Beclin-1 expression, reduced the LC3II/LC3I ratio and promoted p62 expression in GC-1 spg cells in the presence of high glucose, and these effects were all mitigated by rapamycin treatment. Inhibition of PI3K/AKT/mTOR signalling with LY294002 activated autophagy in CLU-overexpressing GC-1 spg cells under high glucose conditions. CLU overexpression repressed autophagy and alleviated testicular damage in diabetic rats, which was also abrogated by LY294002 treatment. CONCLUSIONS: CLU expression is suppressed during diabetes-induced testicular damage, whereas CLU overexpression alleviates diabetes-induced testicular damage by activating PI3K/AKT/mTOR signalling to inhibit autophagy and further repress spermatogenic cell apoptosis.
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Clusterina/fisiología , Diabetes Mellitus Experimental/patología , Testículo , Animales , Apoptosis , Línea Celular , Proliferación Celular , Masculino , Ratones , Proteína Oncogénica v-akt/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Ratas , Ratas Sprague-Dawley , Transducción de Señal , Serina-Treonina Quinasas TOR/metabolismo , Testículo/metabolismo , Testículo/patologíaRESUMEN
PURPOSE: To study the impact of dose distribution on volume-effect parameter and predictive ability of equivalent uniform dose (EUD) model, and to explore the improvements. METHODS AND MATERIALS: The brains of 103 nasopharyngeal carcinoma patients treated with IMRT were segmented according to dose distribution (brain and left/right half-brain for similar distributions but different sizes; V D with different D for different distributions). Predictive ability of EUDV D (EUD of V D ) for radiation-induced brain injury was assessed by receiver operating characteristics curve (ROC) and area under the curve (AUC). The optimal volume-effect parameter a of EUD was selected when AUC was maximal (mAUC). Correlations between mAUC, a and D were analyzed by Pearson correlation analysis. Both mAUC and a in brain and half-brain were compared by using paired samples t-tests. The optimal D V and V D points were selected for a simple comparison. RESULTS: The mAUC of brain/half-brain EUD was 0.819/0.821 and the optimal a value was 21.5/22. When D increased, mAUC of EUDV D increased, while a decreased. The mAUC reached the maximum value when D was 50-55 Gy, and a was always 1 when D ≥55 Gy. The difference of mAUC/a between brain and half-brain was not significant. If a was in range of 1 to 22, AUC of brain/half-brain EUDV55 Gy (0.857-0.830/0.845-0.830) was always larger than that of brain/half-brain EUD (0.681-0.819/0.691-0.821). The AUCs of optimal dose/volume points were 0.801 (brain D2.5 cc), 0.823 (brain V70 Gy), 0.818 (half-brain D1 cc), and 0.827 (half-brain V69 Gy), respectively. Mean dose (equal to EUDV D with a = 1) of high-dose volume (V50 Gy-V60 Gy) was superior to traditional EUD and dose/volume points. CONCLUSION: Volume-effect parameter of EUD is variable and related to dose distribution. EUD with large low-dose volume may not be better than simple dose/volume points. Critical-dose-volume EUD could improve the predictive ability and has an invariant volume-effect parameter. Mean dose may be the case in which critical-dose-volume EUD has the best predictive ability.
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Objective: To investigate the effects of monobromobiphenyl ether (4-BDE) on the expression of γH2AX in the rat testis, and the possible mechanism of 4-BDE affecting the reproductive function of the male rats. METHODS: Twenty-four SD male rats were randomly divided into 4 groups: control and low-, medium- and high-dose 4-BDE, the control rats treated intragastrically with olive oil, and the animals in the latter three groups with 4-BDE at 50, 100 and 200 mg/kg/d, respectively, all for 30 consecutive days. Then all the rats were killed and the testis tissues harvested for HE staining, examination of the apoptosis of the cells by TUNEL and determination of the expressions of H2AX mRNA and γH2AX by q-PCR and Western blot. RESULTS: HE staining manifested occasional reduction or absence of spermatogonial and Sertoli cells in the seminiferous tubules in the medium- and high-dose 4-BDE groups. Compared with the controls, the rats exposed to 4-BDE showed a significant dose-dependent increase in the apoptosis of the testis tissue (P < 0.05), even more significant in the medium- and high-dose 4-BDE groups than in the low-dose group (P < 0.05). There was a dose-effect relationship in the apoptosis index, but with no statistically significant difference between the medium- and high-dose 4-BDE groups (P > 0.05). The results of q-PCR exhibited no statistically significant difference in the expression level of H2AX mRNA either between the control and 4-BDE-exposed rats (P > 0.05) or among the three 4-BDE groups (P > 0.05). The expression of the γH2AX protein was remarkably higher in the 4-BDE groups than in the control (P < 0.05), but not significantly different among the 4-BDE groups (P > 0.05). CONCLUSIONS: Exposure to 4-BDE at 100 or 200 mg/kg/d damages the structure of seminiferous tubules, increases the apoptosis of testicular cells, significantly up-regulates the expression of the γH2AX protein, and consequently increases DNA double-strand breaks (DSB) in the rat testis. The apoptosis of testicular cells may be related to DSB./.
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BACKGROUND: ß-galactosidases are enzymes that are utilized to hydrolyze lactose into galactose and glucose, and are is widely used in the food industry. OBJECTIVE: We describe the recombinant expression of an unstudied, heterodimeric ß-galactosidase originating from Lactobacillus brevis ATCC 367 in Escherichia coli. Furthermore, six different constructs, in which the two protein subunits were fused with different peptide linkers, were also investigated. METHODS: The heterodimeric subunits of the ß-galactosidase were cloned in expressed in various expression constructs, by using either two vectors for the independent expression of each subunit, or using a single Duet vector for the co-expression of the two subunits. RESULTS: The co-expression in two independent expression vectors only resulted in low ß-galactosidase activities, whereas the co-expression in a single Duet vector of the independent and fused subunits increased the ß-galactosidase activity significantly. The recombinant ß-galactosidase showed comparable hydrolyzing properties towards lactose, N-acetyllactosamine, and pNP-ß-D-galactoside. CONCLUSION: The usability of the recombinant L. brevis ß-galactosidase was further demonstrated by the hydrolysis of human, bovine, and goat milk samples. The herein presented fused ß-galactosidase constructs may be of interest for analytical research as well as in food- and biotechnological applications.
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Escherichia coli/enzimología , Lactosa/metabolismo , Levilactobacillus brevis/enzimología , Leche/metabolismo , Fragmentos de Péptidos/metabolismo , beta-Galactosidasa/química , beta-Galactosidasa/metabolismo , Animales , Bovinos , Galactosa/metabolismo , Glucosa/metabolismo , Cabras , Humanos , Hidrólisis , Multimerización de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , beta-Galactosidasa/genéticaRESUMEN
OBJECTIVE: To study the effect of taurine on the reproductive toxicity damage induced by formaldehyde (FM) in adult male rats. METHODS: Forty-eight SD adult male rats were equally randomized into a normal control, an FM poisoning (FMP), a taurine intervention (TI), and an TI+FMP group. The control rats were given normal diet and gavage of saline, the rats of the FMP group treated intraperitoneally with FM at 10 mg/kg qd alt, those of the TI group intragastrically with taurine at 100 mg/kg qd, and those of the TI+FMP group with both FM and taurine at the above doses. After 30 days of treatment, the blood of the abdominal cardinal vein of the rats was extracted for measurement of the levels of serum hormones, the body weight, testis weight and testicular coefficient obtained, the testis tissue subjected to HE staining, and the expressions of Bcl-2 and Bax determined by Western blot. RESULTS: There were no statistically significant differences among the four groups of rats in the body weight, testis weight or testicular coefficient (P > 0.05). The rats in the FMP group showed obviously decreased testicular spermatogenic cells and disordered layers and loose structure of seminiferous tubules, which were basically restored to normal after taurine intervention. Compared with the normal controls, the animals of the TI group exhibited no significant abnormality, but those of the FMP group presented markedly reduced levels of serum T, LH and FSH (P < 0.05), and dramatically increased level of Gonadotropin-releasing hormone (GnRH) (P < 0.01). The levels of serum hormones were all significantly improved (P < 0.05) and that of the apoptotic protein Bax basically returned to normal (P < 0.05) after taurine intervention. CONCLUSIONS: Taurine has a certain protective effect against male reproductive toxicity caused by formaldehyde.
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Formaldehído/toxicidad , Taurina/uso terapéutico , Testículo/efectos de los fármacos , Animales , Masculino , Distribución Aleatoria , Ratas , Ratas Sprague-Dawley , Túbulos Seminíferos/efectos de los fármacos , Túbulos Seminíferos/patología , Testículo/patologíaRESUMEN
PURPOSE: Drug resistance remains a major cause of relapse and therapeutic failure in non-small cell lung cancer (NSCLC). The purpose of this investigation is to explore the relationship between caspase-8 level and chemo-sensitivity, as well as its underlying mechanism in NSCLC cells. METHODS: NSCLC cell line, A549 cells was used to investigate the influence of caspase-8 on the biological behavior in vitro. The abundance of caspase-8 in A549 cells was manipulated by transfection lentivirus containing specific caspase-8 short hairpin RNA (sh-caspase-8) and caspase-8 overexpressed plasmid. Cell viability and the percentage of apoptotic cells was quantified using cell counting kit-8 (CCK-8) assay and flow cytometry following Annexin V-FITC/PI staining, respectively. The formation of acidic vesicle organelles (AVOs) was examined by acridine orange staining and visualized under a fluorescence microscope. The mRNA and protein levels of relative genes were determined by qRT-PCR and western blotting. RESULTS: Our results indicated that cells infected with sh-caspase-8 exhibited high knockdown efficiency. Knockdown of caspase-8 significantly reduced apoptosis of A549 cells. As evidenced by the decreased number of apoptotic cells and the reduction of Bcl-2/bax ratio. Interestingly, caspase-8 knockdown also enhanced autophagy in A549 cells. Additionally, knockdown of caspase-8 reduced the doxorubicin, carboplatin, cisplatin, and etoposide sensitivity towards A549 cells. CONCLUSION: In summary, our results revealed that knockdown of caspase-8 could promote cell growth and autophagy, while reduce chemo-sensitivity and apoptotic cell death. These finding suggest caspase-8 might serve as a potential target to improve the chemo-sensitivity for NSCLC patients in clinical setting.
RESUMEN
PURPOSE: Overweight and obesity are associated with metabolic diseases. However, a subgroup of the overweight/obese population does not present metabolic abnormalities. Hence, there is an urgent need to identify biomarkers that can distinguish different obesity phenotypes and metabolic status. PATIENTS AND METHODS: A total of 98 individuals were divided into three groups: metabolically healthy normal weight (MHNW), metabolically healthy obese (MHO), and metabolically unhealthy obese (MUO). Participants were evaluated for anthropometric and biochemical parameters and serum BMPR1A concentration and miR-503 level. Receiver operating characteristic (ROC) curve analysis and logistic regression analysis were performed. RESULTS: The level of miR-503 was significantly higher in the MHO group compared with that in the MUO group, but no difference was observed between the MHNW and MHO groups. Meanwhile, no significant differences in serum BMPR1A concentration were observed between the three groups. ROC curve analysis showed that miR-503 could be used as a marker to distinguish the MUO from the MHO. Logistic regression analysis suggested that miR-503 was an important related factor associated with an unhealthy metabolic state in overweight/obese subjects. CONCLUSION: miR-503 can be considered as a suitable biomarker to distinguish between the MUO and MHO, which may be a related factor for the incidence of metabolic disorders in overweight/obese subjects.