1.Effect of Biyan Jiedu Capsules on proliferation and apoptosis of nasopharyngeal carcinoma cells based on PI3K/Akt pathway.
Ting LIN ; Yang-Yang TAO ; Ying-Gang TANG ; Ju YUAN ; Hui-Ping DU ; Lin-Yu DENG ; Fang-Liang ZHOU ; Ying-Chun HE
China Journal of Chinese Materia Medica 2025;50(7):1920-1927
To investigate the effects of Biyan Jiedu Capsules on the proliferation and apoptosis of nasopharyngeal carcinoma cells and their molecular mechanism, nasopharyngeal carcinoma cells CNE1 and CNE2 were used. They were divided into control group(30% blank serum medium), low-(10% drug-containing serum + 20% blank serum medium), medium-(20% drug-containing serum + 10% blank serum medium), and high-(30% drug-containing serum medium) concentration group of Biyan Jiedu Capsules according to in vitro experiment. After 24 h of intervention, the effects of Biyan Jiedu Capsules on the proliferation of CNE1 and CNE2 were detected by CCK-8 assay, clonal formation experiment, and EdU staining. The effect of Biyan Jiedu Capsules on apoptosis of CNE1 and CNE2 was detected by flow cytometry. Western blot was used to detect the effect of Biyan Jiedu Capsules on the expression of X-linked apoptosis inhibitor protein(XIAP), survivin, proliferating cell nuclear antigen(PCNA), and PI3K/Akt pathway-related proteins in CNE1 and CNE2. The results showed that compared with the control group, the survival rate of CNE1 and CNE2 in the medium and high concentration groups of Biyan Jiedu Capsules could be decreased in a concentration-dependent way(P<0.05, P<0.01). At the same time, EdU staining and clonal formation experiments showed that the proliferation of CNE1 and CNE2 was significantly inhibited in the medium and high concentration groups of Biyan Jiedu Capsules(P<0.05, P<0.01). Flow cytometry showed that the apoptosis rate of CNE1 and CNE2 was significantly increased in all concentration groups of Biyan Jiedu Capsules(P<0.01), and the apoptosis rate was concentration-dependent. Western blot showed that the expressions of XIAP, survivin, PCNA, p-PI3K, and p-Akt in all concentration groups of Biyan Jiedu Capsules were significantly down-regulated(P<0.05, P<0.01). In conclusion, Biyan Jiedu Capsules can inhibit the proliferation and induce apoptosis of nasopharyngeal carcinoma cells possibly by down-regulating the PI3K/Akt signaling pathway.
Humans
;
Apoptosis/drug effects*
;
Cell Proliferation/drug effects*
;
Nasopharyngeal Carcinoma
;
Nasopharyngeal Neoplasms/physiopathology*
;
Proto-Oncogene Proteins c-akt/genetics*
;
Cell Line, Tumor
;
Drugs, Chinese Herbal/pharmacology*
;
Phosphatidylinositol 3-Kinases/genetics*
;
Signal Transduction/drug effects*
;
Capsules
;
Carcinoma/drug therapy*
2.Predictive biomarkers for immunotherapy in nasopharyngeal carcinoma: from tumor microenvironment to macroenvironment.
Saiwei HUANG ; Yelin LIANG ; Na LIU ; Jun MA
Frontiers of Medicine 2025;19(5):721-742
The introduction of PD-1 blockades to chemotherapy and radiotherapy has shown promising outcomes in patients with nasopharyngeal carcinoma, but anti-PD-1 therapies are only effective in a small proportion of patients, indicating the need for reliable predictive biomarkers of benefit from immunotherapy. Here, we summarized recent advances in immunotherapy for nasopharyngeal carcinoma and studies on potential predictors that correlated with treatment response or long-term survival after immunotherapy, including biomarkers in both the tumor microenvironment and the tumor macroenvironment. Some of these biomarkers have been validated as truly predictive of immunotherapy benefit using cohorts from randomized controlled trials, while others still require further validation of their predictive value. We also summarized the challenges and future directions of biomarker studies, hopefully facilitating the development of predictive biomarkers for immunotherapy that can eventually enter clinical practice.
Humans
;
Tumor Microenvironment/immunology*
;
Nasopharyngeal Carcinoma/immunology*
;
Immunotherapy/methods*
;
Nasopharyngeal Neoplasms/immunology*
;
Biomarkers, Tumor/metabolism*
;
Immune Checkpoint Inhibitors/therapeutic use*
3.miR-34c-3p Inhibits Nasopharyngeal Carcinoma Development via Inhibiting M2 Polarization of Macrophages.
Yu Zi JI ; Yu Jie WANG ; Ji Qing MA ; Zhi Hua YIN ; Fei LIU ; Yan Zi ZANG ; Guang Ke WANG ; Yong TAI
Biomedical and Environmental Sciences 2025;38(2):219-229
OBJECTIVE:
miR-34c-3p is down-regulated in nasopharyngeal carcinoma (NPC). The biological role of miR-34c-3p in NPC and its underlying mechanisms are unknown and were explored in this study.
METHODS:
Flow cytometry and immunohistochemical staining were employed to detect cluster of differentiation 86 (CD86) and cluster of differentiation 206 (CD206) expression; quantitative real-time polymerase chain reaction (qRT-PCR) and western blotting were employed to examine mRNA expression and protein levels; cell counting kit-8 (CCK8) and transwell assays were employed to assess cell proliferation, migration, and invasion; and hematoxylin-eosin (HE) staining was employed to assess pathological changes in tumor tissues.
RESULTS:
Our results revealed that the miR-34c-3p mimic markedly inhibited M2 polarization of macrophages by targeting SLC7A11, and M2 macrophages transfected with the miR-34c-3p mimic inhibited the proliferation, migration, and invasion of NPC cells. The in vivo experiments further confirmed that miR-34c-3p mimics blocked tumor growth and reduced inflammatory infiltration in tumor tissues.
CONCLUSION
This study provides novel insights into the pathogenesis of NPC and a new treatment strategy.
MicroRNAs/metabolism*
;
Nasopharyngeal Carcinoma/genetics*
;
Humans
;
Animals
;
Nasopharyngeal Neoplasms/genetics*
;
Macrophages/physiology*
;
Cell Line, Tumor
;
Mice
;
Cell Proliferation
;
Mice, Inbred BALB C
;
Cell Movement
;
Male
;
Gene Expression Regulation, Neoplastic
;
Mice, Nude
;
Female
4.Clinical observation of free rectus femoris flap for repair of surgical defect in pharyngo-laryngeal malignant tumor.
Wen LI ; Zhe CHEN ; Jiayan WANG ; Xiaojiao LAN ; Zila PURA ; Xiaoxing XIONG ; Liu YANG
Chinese Journal of Reparative and Reconstructive Surgery 2025;39(11):1452-1458
OBJECTIVE:
To explore the feasibility and effectiveness of repairing surgical defect in pharyngo-laryngeal malignant tumor with free rectus femoris flap.
METHODS:
The clinical data of 34 patients with surgical defects in pharyngo-laryngeal malignant tumor who met the selection criteria between July 2014 and August 2024 were retrospectively analyzed. There were 25 males and 9 females, aged 25-82 years, with a median age of 54 years. The disease duration ranged from 2 months to 2 years, with a median of 7 months. The tumor locations included the oropharynx, hypopharynx, cervical esophagus, and larynx. Pathological types included squamous cell carcinoma (29 cases), myoepithelial carcinoma (2 cases), adenoid cystic carcinoma (1 case), and diffuse large B-cell lymphoma (2 cases). TNM staging: 16 cases of T 4N 1M 0, 3 cases of T 4N 2M 0, 3 cases of T 4N 0M 0, 10 cases of T 3N 1M 0, and 2 cases of T 3N 0M 0. The 2017 American Joint Committee on Cancer (AJCC) staging was stage Ⅲ in 2 cases and stage Ⅳ in 32 cases. The blood supply of the proximal rectus femoris muscle was observed by enhanced CT of the lower limb vessels before operation, and the surgical defects ranged from 3.0 cm×2.0 cm to 12.0 cm×8.5 cm. The blood supply and perforators of rectus femoris muscle were explored during operation, and the free rectus femoris flap pedicled with the direct vascular stem of rectus femoris muscle was used to repair the defect. For the patients with pharyngeal fistula or obvious neck swelling after operation, the blood supply of the flap was analyzed by vascular enhanced CT to determine the corresponding strategies of nutritional support, anti-infection, dressing change and drainage. Radiotherapy and chemotherapy were supplemented in 27 patients with lymph node metastasis after operation.
RESULTS:
All the 34 patients were followed up 1-10 years, with an average of 3 years. The flap was found to be necrotic by fibrolaryngoscopy at 1 week after operation in 2 cases, and the incision healed after dressing change and nutritional support, and no reoperation was performed. The flap was in good condition at 1 week after operation in 4 cases, and the signs of gradual necrosis of the flap were found within 1 month after operation, of which 2 cases were healed after dressing change, 1 case was removed the necrotic tissue by reoperation, and 1 case was healed after pectoralis major myocutaneous flap was used to repair the pharyngeal tissue defect. The flaps survived in 28 cases, including 4 cases of pharyngeal fistula, which healed by dressing change. Twenty-two cases achieved satisfactory results in swallowing or phonation. Two patients with total laryngectomy and voice reconstruction underwent reoperation to seal the voice tube because of postoperative aspiration. During the follow-up, 1 case had tracheal stomal recurrence, 2 cases had bone metastasis, and 1 case had bone and lung metastasis.
CONCLUSION
The free rectus femoris flap has good flexibility, the volume of the flap is easy to adjust, and the incision of the donor site is concealed, which is expected to become a new choice for the repair of the surgical defect in pharyngo-laryngeal malignant tumor.
Humans
;
Male
;
Middle Aged
;
Female
;
Aged
;
Adult
;
Plastic Surgery Procedures/methods*
;
Retrospective Studies
;
Laryngeal Neoplasms/pathology*
;
Aged, 80 and over
;
Pharyngeal Neoplasms/pathology*
;
Free Tissue Flaps/blood supply*
;
Quadriceps Muscle/transplantation*
;
Surgical Wound/surgery*
;
Treatment Outcome
5.Evaluation of Clinical Practicability of Hybrid Automatic Treatment Planning for Nasopharyngeal Carcinoma.
Enwei MO ; Lei YU ; Jiyou PENG ; Long YANG ; Jiazhou WANG ; Weigang HU
Chinese Journal of Medical Instrumentation 2025;49(1):55-60
OBJECTIVE:
Automatic planning is a commonly used alternative to manual planning. This study evaluated the clinical performance of automatic plans available in commercial treatment planning systems for nasopharyngeal carcinoma (NPC) treatment by comparing automatic planning with manual planning.
METHODS:
A total of 14 patients with nasopharyngeal carcinoma were enrolled in the study. For each patient, three different sets of clinical goals were used to generate three hybrid automatic plans based on 3D dose distribution prediction and three automatic plans based on script, respectively, which were compared with the manual plans used in clinic.
RESULTS:
The dose coverage performance of the automatic planning based on 3D dose distribution prediction on the planning target volume (PTV) was comparable to that of the manual planning. Automatic planning based on 3D dose prediction achieved the level of manual planning in most organs at risk. However, automatic planning based on scripts did not perform well in the prediction of some organs at risk, especially the parotid gland.
CONCLUSION
The hybrid automatic plan based on 3D dose distribution prediction can reach the level of manual planning and have good robustness with the change of clinical objective.
Humans
;
Nasopharyngeal Neoplasms/radiotherapy*
;
Radiotherapy Planning, Computer-Assisted/methods*
;
Nasopharyngeal Carcinoma
;
Male
;
Female
;
Middle Aged
;
Adult
;
Carcinoma
;
Radiotherapy Dosage
6.Analysis and projection of the disease burden of nasopharyngeal carcinoma in China based on the GBD database.
Yexun SONG ; Xiajing LIU ; Yongquan ZHANG ; Heqing LI
Journal of Central South University(Medical Sciences) 2025;50(4):675-683
OBJECTIVES:
Nasopharyngeal carcinoma is often diagnosed at a late stage due to its concealed location and exhibits marked regional clustering, posing a significant public health challenge in China. This study aims to analyze the disease burden of nasopharyngeal carcinoma in China using the latest 2021 Global Burden of Diseases (GBD) database, providing epidemiological evidence for precise prevention and control of nasopharyngeal carcinoma.
METHODS:
Age-standardized incidence rate (ASIR), mortality rate, and disability-adjusted life year (DALY) rate were used as indicators of disease burden. Stratified analyses were conducted by age, sex, socio-demographic index (SDI), and relevant risk factors. The autoregressive integrated moving average (ARIMA) model and Bayesian age-period-cohort (BAPC) model were employed to project ASIR trends through 2050.
RESULTS:
In 2021, China's age-standardized incidence, mortality, and DALY rates of nasopharyngeal carcinoma were 3.4/100 000, 1.5/100 000, and 48.7/100 000, respectively, all higher than the global average. Across all age groups, Chinese males exhibited higher ASIR, mortality, and DALY rates than females. From 1990 to 2021, the disease burden of nasopharyngeal carcinoma in China decreased gradually with rising SDI. The proportion of nasopharyngeal carcinoma burden attributed to alcohol consumption, smoking, and occupational formaldehyde exposure in China exceeded global levels, especially among males. Projections from both models indicate a rising trend in ASIR for males, females, and the general population in China and globally from 2022 to 2050.
CONCLUSIONS
Over the past 30 years, the disease burden of nasopharyngeal carcinoma in China has decreased with the increasing SDI values but remains higher than the global average. Furthermore, ASIR is projected to increase over the next 30 years. It is imperative for China to enhance healthcare resource allocation for nasopharyngeal carcinoma prevention, diagnosis, and treatment, particularly among high-risk male populations.
Humans
;
China/epidemiology*
;
Male
;
Nasopharyngeal Carcinoma/mortality*
;
Female
;
Middle Aged
;
Nasopharyngeal Neoplasms/mortality*
;
Adult
;
Incidence
;
Global Burden of Disease
;
Disability-Adjusted Life Years
;
Aged
;
Risk Factors
;
Adolescent
;
Databases, Factual
;
Young Adult
;
Cost of Illness
;
Child
;
Bayes Theorem
7.The application of low-temperature plasma minimally invasive therapy in children with pharyngeal neoplasm.
Journal of Clinical Otorhinolaryngology Head and Neck Surgery 2025;39(2):141-146
Objective:Analyze the clinical features in children with pharyngeal neoplasm, and explore the clinical efficacy of low-temperature plasma minimally invasive treatment in children with pharyngeal neoplasm. Methods:A total of 46 pediatric cases of pharyngeal neoplasms admitted to the Otolaryngology Department of Wuhan Children's Hospital from January 2016 to December 2023 were included. There were 23 males and 23 females, with admission ages ranging from 5 days to 9 years and 4 months, and a median age of 2 years and 2 months. The clinical manifestations, examinations, treatments, and outcomes were evaluated. Results:Among the 46 pediatric patients, 21 were admitted with throat wheezing, 7 with masses in the oropharynx or head and neck, 5 with dyspnea, 4 with snoring during sleep as the main symptom, 3 with hoarseness, 2 with sore throat, 2 with swallowing discomfort, and 2 with difficulty feeding as the first symptom. Comorbidities included 5 cases of combined laryngomalacia, 4 of snoring, 4 of congenital heart disease, 4 of severe pneumonia, 3 of myocardial damage, 1 of multiple deformities, and 1 after hematopoietic stem cell transplantation. All patients underwent CT or MRI examination, with 26 cases undergoing local ultrasound examination and 39 undergoing electronic laryngoscopy examination. A total of 19 patients were transferred to the ICU after surgery, all of whom were patients with pharyngeal cysts. All patients underwent low-temperature plasma minimally invasive surgery, and all patients were diagnosed through pathological examination, including 1 case of nasopharyngeal teratoma, 5 of pharyngeal non-Hodgkin's lymphoma, 1 of oropharyngeal rhabdomyosarcoma, 1 of laryngeal perivascular epithelioid cell tumor (PEComa), and 38 of pharyngeal cysts (2 cases of uvula, 2 of oropharynx, 7 of epiglottic valley, 19 of tongue base, 2 of branchial cleft cyst, and 6 of throat). Conclusion:Pharyngeal neoplasms in children are prone to upper airway obstruction, including some rare or malignant tumors. Imaging and pathological diagnosis can assist in early diagnosis. Depending on the nature of the tumor, early detection and treatment can improve the quality of life and survival rate of children. Low-temperature plasma radiofrequency ablation is safe, minimally invasive, precise, and provides a clear field of vision, making it a valuable treatment option for children with pharyngeal neoplasms.
Humans
;
Male
;
Female
;
Child, Preschool
;
Child
;
Infant
;
Pharyngeal Neoplasms/therapy*
;
Minimally Invasive Surgical Procedures/methods*
;
Cold Temperature
;
Treatment Outcome
8.Expression of NFAT5 and IGF1R in nasopharyngeal carcinoma tissues and analysis of clinical characteristics.
Jie YANG ; Qing WANG ; Fusheng LIN ; Lin GAO ; Ran ZHANG ; Xingqian ZHAO ; Xiaojiang LI
Journal of Clinical Otorhinolaryngology Head and Neck Surgery 2025;39(4):333-343
Objective:To investigate the expression of NFAT5 and IGF1R in nasopharyngeal carcinoma tissues and analyze their expression levels in relation to clinical features and prognosis. Methods:From January 1, 2019, to December 31, 2019, 69 cases of nasopharyngeal carcinoma tissues and adjacent non-cancerous tissues were collected from patients treated at Yunnan Cancer Hospital. Immunohistochemistry was employed to detect the expression of NFAT5 and IGF1R in nasopharyngeal carcinoma tissues. The Kaplan-Meier method was used to predict survival time, and the clinicopathological features were evaluated using the log-Rank test. Results:The positive expression rates of NFAT5 and IGF1R in nasopharyngeal carcinoma tissues were 87.0% and 84.5%, respectively. Compared to adjacent normal tissues, the expression levels of NFAT5 and IGF1R in nasopharyngeal carcinoma tissues were significantly increased (P<0.05). Furthermore, the expression of NFAT5 and IGF1R was positively correlated with T stage, N stage, skull base invasion, and cranial nerve palsy (P<0.05). The overexpression of NFAT5 and IGF1R significantly affected the survival rate of patients with nasopharyngeal carcinoma and was negatively correlated with prognosis (P<0.05). Conclusion:In nasopharyngeal carcinoma tissues, overexpression of NFAT5 and IGF1R is observed, which is closely linked to clinical features and patient outcomes. These markers may serve as valuable indicators for predicting the prognosis of nasopharyngeal carcinoma.
Humans
;
Nasopharyngeal Carcinoma/pathology*
;
Nasopharyngeal Neoplasms/metabolism*
;
Prognosis
;
Female
;
Receptor, IGF Type 1/metabolism*
;
Male
;
Transcription Factors/metabolism*
;
Middle Aged
;
Survival Rate
;
Adult
;
Neoplasm Staging
9.Curcumin-loaded nanoparticles reversed radiotherapy-triggered enhancement of MDR1 expression of CNE-2 cells in nasopharyngeal carcinoma.
Guoqing ZENG ; Nan LIAO ; Ning LI ; Gaixia ZHANG ; Yi SU ; Jiangshun SONG
Journal of Clinical Otorhinolaryngology Head and Neck Surgery 2025;39(4):351-356
Objective:This study explored the effect of nanoparticle-encapsulated curcumin on the highly expressed multidrug resistance gene 1 (MDR1) in a human low-differentiated nasopharyngeal carcinoma cell line (CNE2). Methods:Curcumin/chitosan deoxycholic acid nanoparticles were prepared, and the cells were subjected to different treatments: radiotherapy, empty carriers, curcumin, and curcumin-loaded nanoparticles. Cell survival was analyzed using the clonogenic assay, and assessments of apoptosis, MDR1 levels, and miR593 levels were conducted. Results:The cell survival fractions in the curcumin group and the curcumin-loaded nanoparticles group were significantly reduced. Notably, higher apoptosis rates were observed in cells treated with curcumin or curcumin-loaded nanoparticles compared to those that received only radiotherapy. Moreover, a decreased MDR1 level was noted in both the curcumin group and the curcumin-loaded nanoparticles group, with further reduction in MDR1 expression observed in the nanoparticle group (P<0.05). Enhanced expression of miR593 was found in the curcumin group and the curcumin-loaded nanoparticles group, with a relatively higher level in the nanoparticle group (P<0.05). Curcumin encapsulated in nanoparticles exhibited a stronger radiosensitizing effect. The combination of curcumin and radiotherapy effectively inhibited nasopharyngeal carcinoma (NPC) tumor growth, suppressed MDR1 expression, and enhanced miR593 levels. After inhibiting miR593, MDR1 expression increased. The radiosensitizing effect of curcumin-loaded nanoparticles was regulated by miR593 rather than being triggered by MDR1. Conclusion:Curcumin-loaded nanoparticles mediated enhanced expression of miR593, which in turn inhibited the transcription and translation of the MDR1 gene, thereby reducing the radioresistance of NPC and effectively restraining its growth.
Humans
;
Curcumin/pharmacology*
;
Nasopharyngeal Neoplasms/pathology*
;
Nasopharyngeal Carcinoma
;
Nanoparticles
;
Cell Line, Tumor
;
Apoptosis/drug effects*
;
MicroRNAs
;
ATP Binding Cassette Transporter, Subfamily B
;
ATP Binding Cassette Transporter, Subfamily B, Member 1/metabolism*
;
Cell Survival
10.Expression of SLC7A11, GPX4 and ACSL4 in nasopharyngeal carcinoma and their correlation with radiotherapy resistance.
Donghui YAN ; Yanyan ZHENG ; Ningxiang ZENG ; Hongxun GONG ; Yanqiu HUANG ; Maoxin WANG
Journal of Clinical Otorhinolaryngology Head and Neck Surgery 2025;39(5):462-469
Objective:To explore the correlation between ferroptosis-related proteins SLC7A11, GPX4, ACSL4 and the radiosensitivity and prognosis of nasopharyngeal carcinoma. And to investigate the potential of these proteins as molecular markers for predicting the radiosensitivity of nasopharyngeal carcinoma. Methods: A retrospective analysis was conducted on 52 cases of nasopharyngeal carcinoma (nasopharyngeal carcinoma group) and 20 cases of chronic nasopharyngiti s(control group). The relevant clinical data were reviewed, and paraffin-embedded tissue blocks were collected for study. The expressions of SLC7A11, GPX4, and ACSL4 in pathological specimens were detected by immunohistochemical staining. The expression differences of ferroptosis-related proteins between the nasopharyngeal carcinoma group and the control group were analyzed. The nasopharyngeal carcinoma group was further divided based on the protein expression levels into high and low expression subgroups for SLC7A11, GPX4, and ACSL4. Subsequently, a differential analysis of clinical data and survival analysis was conducted for each of these subgroups. Finally, logistic regression analysis was performed to identify the factors influencing radiotherapy resistance in nasopharyngeal carcinoma. Results:①The differential analysis revealed that, compared to the control group, the nasopharyngeal carcinoma group exhibited significantly higher expression of SLC7A11 and GPX4, and lower expression of ACSL4 (P<0.05). ②Notably, the proportion of patients displaying radioresistance was higher in the SLC7A11 and GPX4 high expression groups compared to their respective low expression groups (P<0.05). However, the proportion of radioresistance in the ACSL4 high expression group was lower than that in the ACSL4 low expression group (P<0.05). Survival analysis indicated that the 5-year overall survival rate was lower in the SLC7A11 and GPX4 high expression groups compared to their respective low expression groups(P<0.05). However, the 5-year overall survival rate of the ACSL4 high expression group was higher than that of the ACSL4 low expression group(P<0.05). ③logistic regression analysis showed that SLC7A11 and GPX4 was an independent risk factor for radioresistance in patients with nasopharyngeal carcinoma(P<0.05). Conclusion:Nasopharyngeal carcinoma tissues over-express SLC7A11, GPX4, and under-express ACSL4. Over-expression of SLC7A11 and GPX4 are independent risk factors for radioresistance in patients with nasopharyngeal carcinoma. The inhibition of ferroptosis may be related to the occurrence, progression and radioresistance of nasopharyngeal carcinoma. Detection of the expression of SLC7A11, GPX4, and ACSL4 has guiding significance for the evaluation of radiosensitivity and prognosis of patients with nasopharyngeal carcinoma.
Humans
;
Nasopharyngeal Carcinoma/radiotherapy*
;
Nasopharyngeal Neoplasms/pathology*
;
Coenzyme A Ligases/metabolism*
;
Radiation Tolerance
;
Phospholipid Hydroperoxide Glutathione Peroxidase
;
Amino Acid Transport System y+/metabolism*
;
Prognosis
;
Long-Chain-Fatty-Acid-CoA Ligase
;
Retrospective Studies
;
Ferroptosis
;
Male
;
Female
;
Middle Aged

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