Low concentrations of TNF-α in vitro transform the phenotype of vascular smooth muscle cells and enhance their survival in a three-dimensional culture system.

BACKGROUND: Vascular smooth muscle cells (VSMCs) are commonly used as seed cells in tissue-engineered vascular constructions. However, their variable phenotypes and difficult to control functions pose challenges. This study aimed to overcome these obstacles using a three-dimensional culture system. METHODS: Calf VSMCs were administered tumor necrosis factor-alpha (TNF-α) before culturing in two- and three-dimensional well plates and polyglycolic acid (PGA) scaffolds, respectively. The phenotypic markers of VSMCs were detected by immunofluorescence staining and western blotting, and the proliferation and migration abilities of VSMCs were detected by CCK-8, EDU, cell counting, scratch, and Transwell assays. RESULTS: TNF-α rapidly decreased the contractile phenotypic markers and elevated the synthetic phenotypic markers of VSMCs, as well as markedly increasing the proliferation and migration ability of VSMCs under two- and three-dimensional culture conditions. CONCLUSIONS: TNF-α can rapidly induce a phenotypic shift in VSMCs and change their viability on PGA scaffolds.

Dermoscopic characteristics of membranous aplasia cutis congenita: Report of 56 cases.

BACKGROUND: Membranous aplasia cutis congenita (MACC) presents at birth characterized by oval epidermis defect. Skin lesions with MACC have various clinic manifestations. In recent years, the usefulness of trichoscopy (scalp dermoscopy) has been reported for hair loss diseases. However, the dermoscopic features of MACC were mostly reported by case reports. OBJECTIVES: To summarized the obvious dermoscopic characteristics of MACC. MATERIALS & METHODS: These 56 cases met the clinical diagnostic criteria for MACC without forceps delivery complications or other birth injuries. To find the dermoscopic characteristics of MACC by summarizing 56 infants' dermoscopic pictures. RESULTS: The dermoscopic manifestation of MACC are characterized by hair follicle openings and hair deficiency in the center of skin lesions, translucent epidermis, hair root and hair bulb arranged along the margins of skin lesion. CONCLUSION: The typical dermoscopic characteristics of MACC could help clinicians to early diagnose and differential diagnosis.

High Body Mass Index Worsens Survival in Patients with Esophageal Squamous Cell Carcinoma after Esophagectomy.

AIMS: To investigate the prognostic significance of body mass index (BMI) on the survival of patients with esophageal squamous cell carcinoma (ESCC) after esophagectomy. METHODS: Between 2005 and 2008, 291 patients with ESCC who met the inclusion criteria were included in the study. The BMI cut-off values were as follows: 18.5-23 kg/m2 for normal weight; 23-27.5 kg/m2 for overweight; and ≥27.5 kg/m2 for those with obesity. Univariate and multivariate analyses were performed to identify prognostic factors for long-term survival. RESULTS: Patients were divided into 3 groups: normal weight (n = 138), overweight (n = 103), and obese (n = 50). The median survival time was 56 months. The 5-year overall survival (OS) rates were 40.8, 44.7, and 20.8% for normal weight, overweight, and obese patients respectively (p < 0.05). Multivariate analysis identified BMI as an independent prognostic factor for OS (p < 0.05). For 179 patients without lymph node metastasis, the 5-year OS rates were 46.5, 50.7, and 27.0% for normal weight, overweight, and obese patients respectively (p < 0.05). CONCLUSIONS: A BMI ≥27.5 kg/m2 has a distinctly adverse impact on the long-term survival of ESCC patients after esophagectomy. High BMI is a potential predictor of worse prognosis in ESCC patients, particularly in patients without lymph node metastasis.

Carrageenan maintains the contractile phenotype of vascular smooth muscle cells by increasing macromolecular crowding in vitro.

BACKGROUND: The contractile phenotype of vascular smooth muscle cells (VSMCs) results in good diastolic and contractile capacities, and its altered function is the main pathophysiological basis for diseases such as hypertension. VSMCs exist as a synthetic phenotype in vitro, making it challenging to maintain a contractile phenotype for research. It is widely recognized that the common medium in vitro is significantly less crowded than in the in vivo environment. Additionally, VSMCs have a heightened sense for detecting changes in medium crowding. However, it is unclear whether macromolecular crowding (MMC) helps maintain the VSMCs contractile phenotype. PURPOSE: This study aimed to explore the phenotypic, behavioral and gene expression changes of VSMCs after increasing the crowding degree by adding carrageenan (CR). METHODS: The degree of medium crowding was examined by a dynamic light scattering assay; VSMCs survival and activity were examined by calcein/PI cell activity and toxicity and CCK-8 assays; VSMCs phenotypes and migration were examined by WB and wound healing assays; and gene expression was examined by transcriptomic analysis and RT-qPCR. RESULTS: Notably, 225 µg/mL CR significantly increased the crowding degree of the medium and did not affect cell survival. Simultaneously, CR significantly promoted the contraction phenotypic marker expression in VSMCs, shortened cell length, decreased cell proliferation, and inhibited cell migration. CR significantly altered gene expression in VSMCs. Specifically, 856 genes were upregulated and 1207 genes were downregulated. These alterations primarily affect the cellular ion channel transport, microtubule movement, respiratory metabolism, amino acid transport, and extracellular matrix synthesis. The upregulated genes were primarily involved in the cytoskeleton and contraction processes of VSMCs, whereas the downregulated genes were mainly involved in extracellular matrix synthesis. CONCLUSIONS: The in vitro study showed that VSMCs can maintain the contractile phenotype by sensing changes in the crowding of the culture environment, which can be maintained by adding CR.

Macromolecular Crowding Enhances Matrix Protein Deposition in Tissue-Engineered Vascular Grafts.

Successful in vitro culture of small-diameter tissue-engineered vascular grafts (TEVGs) requires rapid deposition of biomacromolecules secreted by vascular smooth muscle cells in a polyglycolic acid mesh scaffold's three-dimensional (3D) porous environment. However, common media have lower crowding conditions than in vivo tissue fluids. In addition, during the early stages of construction, most of the biomolecules secreted by the cells into the medium are lost, which negatively affects the TEVG culture process. In this study, we propose the use of macromolecular crowding (MMC) to enhance medium crowding to improve the deposition and self-assembly efficiency of major biomolecules in the early stages of TEVG culture. The addition of carrageenan significantly increased the degree of MMC in the culture medium without affecting cell viability, proliferation, and metabolic activity. Protein analysis demonstrated that the deposition of collagen types I and III and fibronectin increased significantly in the cell layers of two-dimensional and 3D smooth muscle cell cultures after the addition of a MMC agent. Collagen type I in the culture medium decreased significantly compared with that in the medium without a MMC agent. Scanning electron microscopy demonstrated that MMC agents considerably enhanced the formation of matrix protein structures during the early stages of 3D culture. Hence, MMC modifies the crowding degree of the culture medium, resulting in the rapid formation of numerous matrix proteins and fiber structures. Impact Statement Small-diameter tissue-engineered vascular grafts (TEVGs) are one of the most promising means of treating cardiovascular diseases; however, the in vitro construction of TEVGs has some limitations, such as slow deposition of extracellular matrix (ECM), long culture period, and poor mechanical properties. We hypothesized that macromolecular crowding can increase the crowding of the culture medium to construct a more bionic microenvironment, which enhances ECM deposition in the medium to the cell layer and reduces collagen loss, accelerating and enhancing TEVG culture and construction in vitro.

Purpura annularis telangiectodes of Majocchi: A case report.

BACKGROUND: Purpura annularis telangiectodes of Majocchi (PATM), also known as Majocchi, is a rare subclass of pigmented purpuric dermatoses. The etiology of PATM is unknown, but it seems more common in children and young women. The skin lesions are mostly symmetrical ring-shaped reddish-brown macules on the lower limbs. CASE SUMMARY: A 9-year-old girl, who has received treated in our department, presented with reddish-brown ring-shaped rash on both lower limbs that had been present for 6 mo. These lesions, red brownish annular or petaloid patches, were mostly found on ankles and lower limber, which do not fade when adding pressure and no feel of infiltration and no atrophy when touching those lesions. Pathological examination showed deposition of hemosiderin in papillary dermis. However, dermoscopy showed the pigmentation in the center as well as the lavender patches on the edge of lesion. The child was thus diagnosed with PATM. After diagnosis, we suggested the patient avoid strenuous exercise. she was given vitamin C tablets for oral and mometasone furoate cream for external use. Follow-up examinations and treatment continue to support the clinical diagnosis to date. CONCLUSION: This is the first report of investigating PATM using dermoscopy, which can differentiate PATM from other diseases due to its unique microscopic feature under dermoscopy. Although PATM is harmless, it still requires long-term follow-up. Moreover, dermoscopy technique can be applied for observation of multi-site lesions and correlated with histopathology. Thus, we believe this approach could be generalized for future diagnosis of PATM.

PRP significantly promotes the adhesion and migration of vascular smooth muscle cells on stent material.

BACKGROUND: The adhesion and survival state of cells on scaffold material is a major problem in tissue-engineered blood vessel (TEBV) culture. Platelet-rich plasma (PRP) contains a large amount of biologically active factors and fibrin, which is expected to play an important role in TEBV culture. PURPOSE: To combine PRP with cells and scaffold material to promote cell adhesion and biological activity on the scaffold material. METHODS: The adhesion status and migration of SMCs under the optimal concentration suitable for SMC growth and the optimal concentration of PRP were examined by scanning electron microscopy, HE staining, CCK-8 assays, qPCR, WB, and other experimental methods and compared with those under the conventional culture (20% FBS); finally, the effect of PRP on the deposition of ECM in vascular tissue engineering culture was verified by three-dimensional culture. RESULTS: PRP at 20% is a suitable concentration for SMCs. Compared with the control group, the 20% PRP group had better migration, and the number of SMC adhesions was significantly higher than that of the control group. In addition, collagen deposition in the experimental group was significantly higher than that in the control group. CONCLUSION: PRP (20%) can promote SMC adhesion, migration, and collagen deposition on the scaffold material.

Neonatal syringocystadenoma papilliferum: A case report.

BACKGROUND: Syringocystadenoma papilliferum (SCAP) represents a rare, noncancerous adnexal tumor predominantly presenting at birth or in early childhood. CASE SUMMARY: In this study, a 35-day-old girl was admitted to Kunming Children's Hospital in October 2019 due to a lesion in the right frontotemporal region since birth. The surface of the lesion was bright red, granular, and papillary and easily bled upon touch, with about 1.5 cm × 4 cm in size. A subcutaneous mass was felt at the base of the lesion, with a size of about 3 cm × 5 cm. Dermatoscopy showed that the skin lesion was lobular and crumby. The lesion center was reddish or white, while the edges were white or yellowish band-like. There were polymorphic vascular structures and white radial streaks in the lesion, with some vascular clusters scattered. Pathological examination showed papilloma-like hyperplasia of the epidermis, with the epidermis partly sinking into the dermis to form several cystic depressions. Combining clinical and histopathological features, the child was diagnosed with SCAP. Follow-up is ongoing, and surgical resection will be performed. CONCLUSION: This was a special clinical manifestation of SCAP, which complements the clinical manifestations of the disease and provides new insights for the diagnosis and differentiation of neonatal skin tumors.

The prevalence of lymph node metastasis for pathological T1 esophageal cancer: a retrospective study of 143 cases.

OBJECTIVE: To evaluate the prevalence, pattern and risk factors of lymph node metastasis (LNM) for pathological T1 (pT1) esophageal cancer (EC). METHODS: The clinical data of 143 cases of pT1 patients who underwent esophagectomy and lymph node dissection during January 2011 and July 2016 were reviewed, including 120 male patients and 23 female patients with a median age of 60 years. The pattern of LNM was analyzed and the risk factors related to LNM were assessed by logistic regression analysis. The nomogram model was used to estimate the individual risk of lymph node metastasis. RESULTS: Of 143 patients with T1 tumors, 25 patients had LNM, and the LNM rate was 17.5%. The LNM rate was 8.0% for T1a tumors, and 22.5% for T1b tumors. The logistic regression analysis showed that the depth of tumor infiltration (P < 0.05), tumor size (P < 0.01), tumor location (P < 0.05), and tumor differentiation (P < 0.01) were independent risk factors related to LNM for T1 EC. These four parameters allowed the compilation of a nomogram to estimate the individual risk of LNM. Tumor differentiation (P < 0.05) was an independent risk factor related to LNM for T1a tumors, and tumor size (P < 0.05) and tumor location (P < 0.05) were independent risk factors related to LNM for T1b tumors. Of 25 patients with LNM, one patient had cervical LNM, 15 patients with thoracic LNM, and 17 patients with abdominal LNM. The relatively highest LNM sites were laryngeal recurrent nerve (n = 8), the left gastric artery (n = 8), right and left cardiac (n = 6) and thoracic paraesophageal (n = 5). CONCLUSIONS: T1 EC has a relatively high LNM rate, and the depth of tumor infiltration, tumor size, tumor location and tumor differentiation are correlated with LNM. The LNM risk and extent must be considered comprehensively in decision-making of a better surgical treatment and lymph node dissection strategy.

Vaccination with allogeneic GM-CSF gene-modified lung cancer cells: antitumor activity comparing with that induced by autologous vaccine.

OBJECTIVE: The aim of this study was to investigate the whole allogeneic (differing tissue-type) tumor cells as vaccine in the mouse lung cancer model. The immunogenic and antitumor activity of allogeneic vaccine was compared with that of autologous cancer cell vaccine. METHODS: C57/BL mice inoculated with Lewis lung cancer (LLC) cells were used as the animal model to test the effects of allogeneic vaccination. LA795 and LLC lung cancer cell lines, which were transfected with the mouse granulocyte-macrophage colony-stimulating factor (GM-CSF) gene, were administered as allogeneic and autologous tumor vaccine, respectively. The irradiated tumor cells were administered as subcutaneous vaccines before the tumor challenge. The immunity of cancer vaccine was tested by mouse interferon-gamma (IFN-gamma) enzyme-linked immunospot (ELISPOT) lactate dehydrogenase (LDH) assays. The serum level of IFN-gamma and interleukin (IL)-4 was tested using the enzyme-linked immunosorbent assay method. RESULTS: Prophylactic vaccination with allogeneic LA795 cells protected against the LLC tumor challenge in C57/BL. The tumor growth was inhibited and the survival was accordingly prolonged. The cytotoxicity of the spleen cells or the purified CD(8)(+) T-cells against LLC cells in the mice immunized with either the autologous or allogeneic cancer cell vaccine was significantly increased, relative to that of the control, untreated group (p<0.05). ELISPOT IFN-gamma assays showed that spleen cells from mice immunized with LA795 cells could be activated after coculture with irradiated LLC cells. In addition, the serum level of Th1-king cytokine IFN-gamma significantly increased after vaccination; however, no statistically difference was found in Th2-kind cytokine IL-4. CONCLUSIONS: The allogeneic cancer vaccine could induce immune responses and protection against lung cancer, which had no significant difference with that of autologous vaccine.

[The anti-tumor activity of GM-CSF-modified lung cancer cell vaccine and its synergism in combination with chemotherapy].

OBJECTIVE: To investigate the anti-tumor effects and mechanism of tumor vaccines and whether chemotherapeutic agents administered prior to immunotherapy could augment the efficacy of the vaccines. METHODS: C57/BL mice inoculated with Lewis lung cancer cells were used as tumor models. Granulocyte-macrophage colony-stimulating factor (GM-CSF) gene modified LA795 and Lewis lung cancer cell lines were administered as allogeneic and autologous tumor vaccines, respectively. After Lewis cells (1 x 10(7)) inoculation, the mice received irradiated GM-CSF secreting cancer vaccine solely or in combination with carboplatin. The survival of the mice was observed. The cytotoxicity of spleen cells or purified CD8(+) cells was analyzed by lactate dehydrogenase (LDH) assay. Serum level of IL-4 and IFN-gamma was detected using ELISA method. RESULTS: The cytotoxicity of the spleen cells or purified CD8(+) T cells against Lewis cells in the mice immunized with cancer cell vaccine was significantly increased, relative to that of the control, untreated group (P < 0.05). Serum level of Th1-type cytokine IFN-gamma was increased after vaccination, whereas Th2-type cytokine IL-4 showed no significant change. The GM-CSF secreting cancer cell vaccine had no significant influence on the survival of the mice with established heavy tumor burden. The combination of chemotherapy and cancer vaccine could statistically prolong the survival time; whereas any method itself had no significant effect. CONCLUSION: The GM-CSF secreting cancer cell vaccine can induce immune responses. The chemotherapeutic agents may be beneficial to enhance the anti-tumor activity of cancer vaccine.

Lymph node dissection for Siewert II esophagogastric junction adenocarcinoma: A retrospective study of 3 surgical procedures.

The present study was aimed to investigate the application of right thansthoracic Ivor-Lewis (IL), left transthoracic (LTT), and left thoracoabdominal (LTA) approach in Siewert type II adenocarcinoma of esophagogastric junction (AEG).The data of 196 patients with Siewert type II AEG received surgical resection in our cancer center between January 2014 and April 2016 was retrospectively analyzed. Finally, 136 patients met the inclusion criteria were enrolled in the study and divided into the IL (47 cases), LTT (51 cases), and LTA group (38 cases). Clinical and short-term treatment effects were compared among the 3 groups.The patients with weight loss, diabetes, and heart disease increased in the LTT group (P = 0.054, P = 0.075, and P = 0.063, respectively). Operation time was significantly longest in the IL group (P < 0.001), but the amount of bleeding and tumor size did not significantly differ among the 3 groups (P = 0.176 and P = 0.228, respectively). The IL group had the significantly longest proximal surgical margin (P < 0.001) and most number of total (P < 0.001) and thoracic lymph nodes (P < 0.001) dissected. Both the IL and LTA groups had more abdominal lymph nodes dissected than the LTT group (P < 0.001). In general, the IL and LTT groups had the highest dissection rates of every station of thoracic (P < 0.05) and lower mediastinal lymph nodes (P < 0.05), respectively. The dissection rate of the paracardial, left gastric artery, and gastric lesser curvature lymph nodes did not differ significantly among the 3 groups (P > 0.05), but the dissection rate of the hepatic artery, splenic artery, and celiac trunk lymph nodes was significantly highest in the IL group (P < 0.05). Postoperative hospital stay, perioperative complications, and mortality did not differ significantly among the 3 groups (P > 0.05).Compared with the traditional left transthoracic approach, the Ivor-Lewis approach did not increase the perioperative mortality and complication rates in Siewert type II AEG, but obtained satisfactory length of the proximal surgical margin, and was better than the left transthoracic approach in thoracic and abdominal lymph node dissection. However, the advantages of Ivor-Lewis procedure requires further follow-up and validation through prospective randomized controlled trials.

Metastatic to negative lymph node ratio demonstrates significant prognostic value in patients with esophageal squamous cell carcinoma after esophagectomy.

AIMS: The prognostic value of metastatic lymph node ratio (LNR) has been reported in some studies; however, there is no report on the prognostic significance of metastatic to negative lymph node ratio (MNLNR) in cancer patients. The aim of this study was to compare the prognostic value of pN, LNR and MNLNR on the survival of patients with esophageal squamous cell carcinoma (ESCC) after esophagectomy. METHODS: The data of 290 patients with ESCC after curative esophagectomy was retrospectively analyzed. The optimal cut-point for LNR and MNLNR were set as 0, 01-0.2, and >0.2. Univariate and multivariate analyses were performed to identify prognostic factors for overall survival (OS). RESULTS: Patients classified as LNR 0, 0.01-0.20, and 0.21-1.0, the observed 5-year OS rates were 46.6%, 26.0%, and 11.6%, respectively (P = 0.000). Patients classified as MNLNR 0, 0.01-0.20, and >0.2, the observed 5-year OS rates were 46.6%, 31.2%, and 7.4%, respectively, respectively (P = 0.000). The pN stage, LNR or MNLNR category was confirmed as a significant independent prognostic factor, respectively (P = 0.032, P = 0.011 and P = 0.003, respectively); However, only the MNLNR category (P = 0.003) remained as a significant prognostic factor when the pN stage, LNR and MNLNR category simultaneously included in the multivariate analysis models. CONCLUSIONS: The MNLNR was recognized as an independent prognostic factor in ESCC patients after curative esophagectomy. In addition, MNLNR showed better prognostic value than pN stage and LNR category.

Is tumor length a prognostic indicator for esophageal squamous cell carcinoma? A single larger study among Chinese patients.

OBJECTIVE: In esophageal cancer, depth of wall penetration, reflected by T classification, represents the most important prognostic variable. Our study aimed to investigate the impact of tumor length, measured as the longitudinal length, on the outcome of esophageal squamous cell carcinoma (ESCC) patients. METHODS: The survival data of 362 ESCC patients who underwent surgical resection as the primary treatment between 1999 and 2007 were collected retrospectively. Receiver-operator characteristic analysis was applied to identify the optimal cut-off values. RESULTS: 4.0 cm was identified as the optimal cut-off value within the whole group. Tumor length greater than 4.0 cm was associated with increasing T stage (P=0.001), N stage (P=0.046), and tumor differentiation (P=0.033). Univariate analysis and multivariate analysis both found that tumor length greater than 4.0 cm was associated with worse overall survival compared with shorter tumors (P<0.001). It appeared to have a greater impact on N0-N1 (P<0.001, P=0.026, respectively) than N2-N3 and appeared to have a higher impact on the lower-stage patients than the higher-stage patients. CONCLUSIONS: Tumor length proved to be an independent prognostic parameter for ESCC patients, especially for node-negative and lower-stage patients. More attention should be paid to its role in the management of ESCC.

Association of Kruppel-like factor 4 expression with the prognosis of esophageal squamous cell carcinoma patients.

OBJECTIVE: To investigate the association of Kruppel-like factor 4 (KLF4) expressions with the prognosis of esophageal squamous cell carcinoma (SCC) patients. METHODS: Ninety-eight cases of esophageal carcinoma patients were enrolled. The expression of KLF4 in the esophageal SCC and normal esophageal mucosa tissues were examined by immunohistochemistry. The correlations between the expression of KLF4 protein and patients' clinical characteristics and prognosis were analyzed. RESULTS: We observed higher expressed KLF4 in normal esophageal mucosa tissues than esophageal SCC tissues, with positive rate of 82.7% (81/98) and 43.8% (43/98) respectively. In patients with lymphatic metastasis, the positive rate of KLF4 was 24.4% (10/41), whereas it was 57.9% (33/57) in patients without lymphatic metastasis, and the difference was significant (x(2) = 10.871, P = 0.001). The positive rates of KLF4 were 62.5% (5/8), 53.1% (26/49) and 29.3% (12/41) in stage I, II and III patients, respectively. There were no correlations between the expression of KLF4 and gender, age, tumor size, location, differentiation grade and infiltration depth. The 5-year survival rates and median survival times were 48.8% and 25.5%, and 55 and 26 months for the patients with KLF4 positive and negative expression, respectively. There were significant differences between the patients with KLF4 positive expression and negative expression in the 5-year survival rates and median survival times (x(2) = 5.747 and 4.493, P = 0.017 and 0.034). CONCLUSION: KLF4 might act as a tumor suppressor in esophageal SCC and the expression status of KLF4 could be considered as a prognosis predictor for esophageal SCC patients.