Efficient biosynthesis of γ-aminobutyric acid by rationally engineering the catalytic pH range of a glutamate decarboxylase from Lactobacillus plantarum / 生物工程学报

γ-aminobutyric acid can be produced by a one-step enzymatic reaction catalyzed by glutamic acid decarboxylase. The reaction system is simple and environmentally friendly. However, the majority of GAD enzymes catalyze the reaction under acidic pH at a relatively narrow range. Thus, inorganic salts are usually needed to maintain the optimal catalytic environment, which adds additional components to the reaction system. In addition, the pH of solution will gradually rise along with the production of γ-aminobutyric acid, which is not conducive for GAD to function continuously. In this study, we cloned the glutamate decarboxylase LpGAD from a Lactobacillus plantarum capable of efficiently producing γ-aminobutyric acid, and rationally engineered the catalytic pH range of LpGAD based on surface charge. A triple point mutant LpGADS24R/D88R/Y309K was obtained from different combinations of 9 point mutations. The enzyme activity at pH 6.0 was 1.68 times of that of the wild type, suggesting the catalytic pH range of the mutant was widened, and the possible mechanism underpinning this increase was discussed through kinetic simulation. Furthermore, we overexpressed the Lpgad and LpgadS24R/D88R/Y309K genes in Corynebacterium glutamicum E01 and optimized the transformation conditions. An optimized whole cell transformation process was conducted under 40 ℃, cell mass (OD600) 20, 100 g/L l-glutamic acid substrate and 100 μmol/L pyridoxal 5-phosphate. The γ-aminobutyric acid titer of the recombinant strain reached 402.8 g/L in a fed-batch reaction carried out in a 5 L fermenter without adjusting pH, which was 1.63 times higher than that of the control. This study expanded the catalytic pH range of and increased the enzyme activity of LpGAD. The improved production efficiency of γ-aminobutyric acid may facilitate its large-scale production.

Protection of intestinal immune barrier against ischemia/reperfusion injury in a swine model using anisodamine hydrobromide

Abstract Intestinal ischemia/reperfusion (I/R) causes barrier impairment and bacterial influx. This study explored the protective effects of anisodamine hydrobromide (AH) on intestinal I/R injury caused by cardiopulmonary resuscitation (CPR) after cardiac arrest (CA). After successful CPR, minipigs were randomly divided into two groups (n = 8): saline and AH (4 mg/kg), and then treated with saline or AH via central venous injection, respectively. The same procedures without ventricular fibrillation initiation were conducted in the Sham group (n = 8). Levels of interferon gamma (IFN-γ) and interleukin 4 (IL-4) were measured at different time points (0, 0.5, 1, 2, 4, and 6 h) in serum and 6 h in gut associated lymphoid tissues (GALTs) after the return of spontaneous circulation (ROSC) to evaluate changes in the proportion of T-helper type 1 (Th1) and T-helper type 2 (Th2). Moreover, the positive culture rates of GALTs were examined to evaluate bacterial translocation. AH treatment markedly alleviated aberrant arterial blood gas and hemodynamics as well as intestinal macroscopic and morphological changes after CPR. Moreover, AH treatment significantly increased IFN-γ and decreased IL-4 in both serum and GALTs. Furthermore, AH treatment dramatically decreased positive bacterial growth in GALTs. AH treatment mitigated immunosuppression caused by intestinal I/R and protected the intestinal immune barrier against bacterial translocation, thereby reducing the risk of secondary intestinal infection

Significance of lysophosphatidic acid receptor 6 in the large-cell transformation of mycosis fungoides and its effect on the proliferation and apoptosis of cutaneous T-cell lymphoma cells / 中华皮肤科杂志

Objective:To determine lysophosphatidic acid receptor 6 (LPAR6) expression in patients with mycosis fungoides (MF) , a variant of cutaneous T-cell lymphoma (CTCL) , and to investigate its role and mechanism of action in the development and prognosis of CTCL.Methods:A total of 110 patients with confirmed MF were collected from Department of Dermatology, Peking University First Hospital from 2011 to 2020, including 24 with large-cell transformation (LCT) and 25 with non-large cell transformation (NLCT) in the discovery cohort, and 24 with LCT and 37 with NLCT in the validation cohort. RNA sequencing and RT-PCR were conducted to determine the LPAR6 expression in patients in the discovery cohort and validation cohort respectively. LPAR6 expression was compared between patients with LCT and those with NLCT, and its effect on the prognosis of patients was evaluated. Two LPAR6-overexpressing CTCL cell lines MyLa and Sz4 were constructed to evaluate the effect of LPAR6 overexpression on proliferative activity of MyLa and Sz4 cells, with the cells normally expressing LPAR6 as the control group; after the treatment with LPAR6-related ligand lysophosphatidic acid (LPA) , 2S-OMPT, adenosine triphosphate (ATP) or adenosine (ADO) , the effects of LPAR6 activation on the proliferative activity and apoptosis of LPAR6-overexpressing MyLa and Sz4 cells were evaluated by the MTS method and flow cytometry respectively. Log-rank test was used for prognostic analysis, and t test or Mann-Whitney U test was used for comparisons between two groups. Results:As RNA sequencing showed, LPAR6 was one of the significantly underexpressed genes in the LCT group in the discovery cohort; in the validation cohort, LPAR6 expression (median[ Q1, Q3]) was significantly lower in the LCT group (204.90[81.90, 512.70]) than in the NLCT group (809.40[417.50, 1 829.20], U= 242.00, P= 0.002) ; in the two cohorts, the underexpression of LPAR6 was significantly associated with increased risk of poor prognosis (both P < 0.01) . Cell proliferation assay showed no significant difference in the proliferative activity of MyLa or Sz4 cells between the LPAR6 overexpression group and control group at 0, 24, 48 and 72 hours during the experiment (all P > 0.05) ; 48 hours after activation of LPAR6 by LPA, 2S-OMPT, ATP and ADO in MyLa cells, the LPAR6 overexpression group showed significantly decreased cellular proliferative activity (1.38 ± 0.01, 1.04 ± 0.01, 1.09 ± 0.03, 1.23 ± 0.01, respectively) compared the control group (1.73 ± 0.04, 1.23 ± 0.01, 1.24 ± 0.01, 1.42 ± 0.03, t= 30.33, 18.38, 4.78, 5.75, respectively, all P < 0.05) , but significantly increased cell apoptosis rate (17.93% ± 0.88%, 17.75% ± 0.35%, 23.97% ± 0.57%, 31.44% ± 0.34%, respectively) compared the control group (3.98% ± 0.03%, 7.81% ± 0.59%, 11.95% ± 0.85%, 12.02% ± 0.48%, t= 15.93, 14.49, 11.74, 33.01, respectively, all P < 0.05) ; 48 hours after activation of LPAR6 by 2S-OMPT and ADO in Sz4 cells, compared with the control group, the LPAR6 overexpression group also showed significantly decreased cellular proliferative activity (2S-OMPT: 1.29 ± 0.04 vs. 1.48 ± 0.01; ADO: 1.27 ± 0.01 vs. 1.51 ± 0.02; both P < 0.05) , but significantly increased cell apoptosis rate (2S-OMPT: 41.70% ± 0.70% vs. 29.35% ± 0.55%; ADO: 37.05% ± 0.15% vs. 24.60% ± 1.00%; both P < 0.05) . Conclusions:LPAR6 was underexpressed in the patients with LCT, and its underexpression was significantly associated with increased risk of poor prognosis. In vitro activation of LPAR6 could inhibit the proliferation of CTCL cells and promote their apoptosis, suggesting that the decrease of LPAR6 expression may be one of the important mechanisms underlying disease progression in patients with LCT.

Clinicopathological and immunophenotypic analysis of 24 cases of transformed mycosis fungoides / 中华皮肤科杂志

Objective:To investigate clinicopathological features and prognosis of transformed mycosis fungoides (TMF) .Methods:A retrospective analysis was performed on clinicopathological data collected from 24 patients with TMF, as well as on flow cytometry results of 16 peripheral blood samples obtained from 11 of the 24 patients, who visited Hospital of Dermatology, Chinese Academy of Medical Sciences between 2014 and 2020.Results:Among the 24 patients, 11 were males and 13 were females. Their average age at diagnosis of TMF was 50.0 years (range: 18 - 77 years), and patients with early-stage TMF (9 cases) and tumor-stage TMF (15 cases) were aged 44.8 and 52.6 years on average, respectively. The average time interval from diagnosis of MF to large cell transformation was 3.7 years, and 8 patients were diagnosed with TMF at the initial visit. Histopathologically, large cells infiltrated in a diffuse pattern in 20 cases, as well as in a multifocal pattern in 4, and the proportion of large cells in 7 cases was greater than 75%. Immunohistochemically, 18 patients showed positive staining for CD30, and the proportion of CD30-positive large cells was greater than 75% in 9; negative staining for CD30 was observed in 6. Flow cytometry of 16 peripheral blood samples showed the presence of cell subsets expressing clonal T cell receptor (TCR) -vβ in 2 of 4 patients with early-stage TMF and 10 of 12 with tumor-stage TMF, and tumor cells with higher forward scatter than normal lymphocytes were detected in 16 samples. During the follow-up, among the patients with early-stage TMF, 3 progressed to tumor-stage TMF 3.3 years on average after large cell transformation, 1 progressed to erythrodermic MF in stage IIIA, and the other 4 still showed an indolent course; among the patients with tumor-stage TMF, 1 progressed to stage-IV TMF, and 5 died 3.3 (1.5 - 6) years after large cell transformation.Conclusion:Large cell transformation may occur in patients with MF in any stage, some patients have poor prognosis, so close follow-up is needed for patients with TMF.

Identificação de alterações bioquímicas em células epiteliais bronquiais humanas normais expostas a fumarato / Identification of biochemical changes in normal human bronchial epitelial cells exposed to fumarate

A reprogramação metabólica de células do câncer é apontada como uma característica essencial para o desenvolvimento da doença (cancer hallmark). Estudos mostram que mutações na enzima fumarato hidratase levam ao aumento da concentração intra e extracelular de fumarato, o que ocorre paralelamente à indução da transformação maligna. Neste trabalho, a fim de entender se o excesso de fumarato extracelular pode propiciar a transformação de células normais, foram quantificados alguns endpoints relacionados aos efeitos do fumarato e à transformação maligna, além de alterações metabólicas em células imortalizadas de epitélio brônquico humano normal (BEAS-2B) expostas ao fumarato. Uma vez que fumarato nas concentrações de 0,1 a 10 mM ao longo de 144 h não foi citotóxico, foram selecionadas as concentrações de 1 mM, 5 mM e 10 mM para as incubações. Fumarato induziu a formação de colônias em soft-agar após o período de sete dias (168 h) de exposição, o que indica a indução de transformação celular. Fumarato é um oncometabólito que inibe enzimas que dependem de α-cetoglutarato como co-substrato, dentre as quais as enzimas ten eleven translocation (TET) que catalisam a formação de 5-hidroximetilcitosina (5-hmC) a partir de 5-metilcitosina (5-mC), o primeiro passo da sequência de reações que levam à desmetilação do DNA. Os níveis totais de 5-hmC estavam diminuídos no DNA das células expostas. Colônias retiradas do soft-agar (controle, 1, 5 e 10 mM de fumarato) foram cultivadas e, após 90 dias em cultura, as células foram submetidas ao ensaio de invasão e migração em câmara de Boyden (transwell), tendo sido observada maior capacidade de migração/invasão das células anteriormente expostas ao fumarato. Foi observada indução de estresse redox nas células expostas ao fumarato. A partir da quantificação de metabólitos intracelulares por HPLC-ESI-MS/MS e HPLC-ESI-Q-TOF, verificamos que as células BEAS-2B absorveram o fumarato adicionado ao meio de cultura, o qual foi convertido intracelularmente a malato, aspartato, argininosuccinato, citrato, succinato e glutamato. O oncometabólito 2-L-hidroxiglutarato foi detectado em níveis aumentados nas células expostas a fumarato, assim como adenosina, enquanto que NAD+ e NADP+ apareceram diminuídos. As alterações metabólicas na presença de fumarato contribuíram para a manutenção do balanço energético das células, ou mesmo para um saldo positivo de energia. A exposição das células a [13C4]fumarato permitiu a análise do fluxo inicial do fumarato absorvido pelas células. A partir dessa análise verificamos que o fumarato absorvido entra no ciclo de Krebs, gerando malato, que é em grande parte desviado para reações externas ao ciclo, como a geração de aspartato e argininosuccinato. Citrato proveniente das reações de [13C4]fumarato no ciclo de Krebs foi detectado em níveis inferiores aos endógenos. O uso de [13C4]fumarato permitiu a visualização da geração de [13C4]succinato, que tem como possível fonte a atividade reversa da succinato desidrogenase. Verificamos também a geração de [13C3]glutamato. Supõe-se que as alterações metabólicas induzidas pelo fumarato absorvido pelas células BEAS-2B contribuam para a modulação da expressão de genes e da atividade de proteínas que favorecem o processo tumorigênico

The metabolic reprogramming of cancer cells is identified as an essential feature for the development of the disease (a cancer hallmark). Studies show that mutations in the enzyme fumarate hydratase lead to increased intra- and extracellular fumarate concentration, which occurs in parallel with the induction of malignant transformation. In this work, in order to understand if excess extracellular fumarate can lead to the transformation of normal cells, some endpoints related to the effects of fumarate and malignant transformation were quantified, as well as metabolic alterations in the immortalized normal human bronchial epithelial cell line BEAS-2B exposed to fumarate. Since fumarate at concentrations from 0.1 to 10 mM over 144 h was not cytotoxic, the concentrations of 1 mM, 5 mM and 10 mM were selected for incubations. Fumarate induced colony formation in soft agar after the seven day (168 h) exposure period, which indicates the induction of cell transformation. Fumarate is an oncometabolite that inhibits α-ketoglutarate-dependent enzymes, among which are ten eleven translocation (TET) enzymes that catalyze the formation of 5-hydroxymethylcytosine (5-hmC) from 5-methylcytosine (5-mC), the first step in the sequence of reactions leading to DNA demethylation. Total 5-hmC levels were decreased in the DNA of exposed cells. Colonies removed from the soft-agar (control, 1, 5 and 10 mM fumarate) were cultured and after 90 days in culture the cells were subjected to the Boyden chamber (transwell) invasion and migration assay, and a greater capacity for migration/invasion of cells previously exposed to fumarate was observed. Redox stress induction was observed in cells exposed to fumarate. From the quantification of intracellular metabolites by HPLC-ESI-MS/MS and HPLC-ESI-Q-TOF, we found that BEAS-2B cells absorbed the fumarate added to the culture medium, which was intracellularly converted to malate, aspartate, argininosuccinate, citrate, succinate and glutamate. The oncometabolite 2-L-hydroxyglutarate was detected at increased levels in cells exposed to fumarate, as well as adenosine, while NAD+ and NADP+ appeared decreased. Metabolic changes in the presence of fumarate contributed to the maintenance of the energy balance of the cells, or even to a positive energy balance. Exposure of cells to [13C4]fumarate allowed the analysis of the initial flow of the fumarate absorbed by the cells. From this analysis we found that the absorbed fumarate entered the Krebs cycle, generating malate, which was largely diverted to reactions outside the cycle, such as the generation of aspartate and argininosuccinate. Citrate from the reactions of [13C4]fumarate in the Krebs cycle was detected at levels lower than endogenous. The use of [13C4]fumarate allowed the detection of [13C4]succinate, which has as its possible source the reverse activity of succinate dehydrogenase. We also observed the generation of [13C3]glutamate. The metabolic changes induced by the absorbed fumarate are supposed to contribute to the modulation of gene expression and protein activity that favor the tumorigenic process

Rational design of the C-terminal Loop region of leucine dehydrogenase and cascade biosynthesis L-2-aminobutyric acid / 生物工程学报

Leucine dehydrogenase (LDH) is the key rate-limiting enzyme in the production of L-2-aminobutyric acid (L-2-ABA). In this study, we modified the C-terminal Loop region of this enzyme to improve the specific enzyme activity and stability for efficient synthesis of L-2-ABA. Using molecular dynamics simulation of LDH, we analyzed the change of root mean square fluctuation (RMSF), rationally designed the Loop region with greatly fluctuated RMSF, and obtained a mutant EsLDHD2 with a specific enzyme activity 23.2% higher than that of the wild type. Since the rate of the threonine deaminase-catalyzed reaction converting L-threonine into 2-ketobutyrate was so fast, the multi-enzyme cascade catalysis system became unbalanced. Therefore, the LDH and the formate dehydrogenase were double copied in a new construct E. coli BL21/pACYCDuet-RM. Compared with E. coli BL21/pACYCDuet-RO, the molar conversion rate of L-2-ABA increased by 74.6%. The whole cell biotransformation conditions were optimized and the optimal pH, temperature and substrate concentration were 7.5, 35 °C and 80 g/L, respectively. Under these conditions, the molar conversion rate was higher than 99%. Finally, 80 g and 40 g L-threonine were consecutively fed into a 1 L reaction mixture under the optimal conversion conditions, producing 97.9 g L-2-ABA. Thus, this strategy provides a green and efficient synthesis of L-2-ABA, and has great industrial application potential.

Research progress in cell transformation test and its application in cigarette smoke evaluation / 公共卫生与预防医学

In order to understand the current development of cell transformation assay (CTA) and its application in the evaluation of cigarette smoke carcinogenesis, the relevant literatures were analyzed and combed from these two aspects. CTA can evaluate the carcinogenicity of various genotoxic and non-genotoxic carcinogens in a short period of time, and has a strong consistency with the results of animal carcinogenic test. After malignant transformation, the cells show changes in cell morphology, immortalization of cells, disappearance of cell-cell contact inhibition, and the ability to form tumors when injected into animals. The identification methods of transformed cells include transformed cell focus count, agglutination test, soft agar culture and inoculation of nude mice, etc. At present, BALB/c 3T3 cells, Bhas 42 cells and SHE cells are the most widely used cells for CTA. Cigarette smoke is a complex aerosol containing a variety of non-genetic carcinogenic chemicals. Cell transformation tests are often used as an in vitro alternative method to evaluate the carcinogenic effects of cigarette smoke, which is different from the short-term genetic toxicity test. It simulates the long-term state of human smoking induced malignant transformation of cells, through the long-term exposure of cells for several decades, which is closer to the occurrence of cancer caused by human smoking. Therefore, CTA can evaluate the carcinogenicity of cigarette smoke and other tobacco products.

Study on the Effects of Calpeptin on Estrogen-induced Transformation and Stemness Markers Expression of Mammary Epithelial Cells MCF- 10A Based on Calpain-ERK Signaling Pathway / 中国药房

OBJECTIVE：To study the effects of Calpeptin inhibitor Calpeptin on the transformation and stemness markers expression induced by estradiol（E2），and to investigate its mechanism. METHODS ：Taking human mammary epithelial cells MCF-10A as research object ，transformed cells were induced by E 2 treatment. Cells were divided into control group （0.1％DMSO）， E2-transformed group （50 nmol/L），E2-transformed+Calpeptin group （50 nmol/L E 2+1 μmol/L Calpeptin），then continuously treated with corresponding drug-containing culture medium for 15 generations. Then ，MTT assay was used to determine the proliferation rate of cells （24，48 h）；plate colony test was used to detect the Clone formation rate of cells ；the number of sphere-forming cells was measured by suspension spheroidization test ；mRNA expressions of stemness marker （CD44，Nanog，OCT4）and extracellular sigal-regulated kinase （ERK）were detected by RT-qPCR ，and protein expressions of CD 44，Nanog，OCT4 ，ERK and p-ERK were detected by Western blotting assay. Another E 2-transformed cells were divided into control group （0.1％DMSO）and U0126 （ERK inhibitor ）group（10 μmol/L）. Clone formation rate ，the number of sphere-forming ，protein expressions of CD 44，Nanog， OCT4，ERK and p-ERK were determined with above methods ，and to validate the relationship of ERK inhibition with transformed cell behavior and the expression of stemness markers. RESULTS ：Compared with control group ，proliferation rate and clone formation rate of E 2 transformed group were increased significantly （P＜0.01），and the number of sphere-forming was increased significantly（P＜0.01）；mRNA expression levels of CD 44，Nanog，OCT4，ERK and protein expression levels of CD 44，Nanog， OCT4 and p-ERK in cells were increased significantly （P＜0.01）. Compared with E 2-transformed group ，proliferation rate （24，48 h）and clone formation rate of E 2-transformed + Calpeptin group were decreased significantly （P＜0.01），and the number of sphere-forming was decreased significantly （P＜0.05）；mRNA expression levels of CD 44，Nanog，OCT4 ，ERK and protein expression levels of CD 44，Nanog，OCT4，p-ERK in cells were decreased significantly （P＜0.05 or P＜0.01）. After treated with ERK inhibitor U 0126，clone formation rate of E 2-transformed cells ，the number of sphere-forming ，protein expression levels of CD44，Nanog，OCT4 and p-ERK were increased significantly （P＜0.05 or P＜0.01）. CONCLUSIONS ：Calpeptin can inhibit the transformation and the expression of stemness markers of human mammary epithelial cells MCF- 10A，and the mechanism of it may be associated with inhibiting the activation of Calpain-ERK signaling pathway.

Efficacy and safety of beta carotones in treatment of oral leukoplakia: systematic review and meta-analysis / Eficacia y seguridad de las betacarotonas en el tratamiento de la leucoplasia oral: revisión sistemática y metanálisis

Objectives: A systematic review was conducted to evaluate effectiveness and safety of beta carotenes for the treatment of oral leukoplakia regarding clinical resolution and prevention of malignant transformation. Material and Methods: The systematic search was conducted in three electronic databases and the study's selection was performed according to pre-set eligibility criteria. Four studies evaluating the efficacy of beta carotenes in oral leukoplakia compared to placebo were included in the review; three of which were assigned for quantitative analysis. Data were extracted, tabulated, quality assessed and statistically analyzed. Results: The meta-analysis revealed that when comparing clinical resolution the beta carotene group favored was favored compared to placebo, with statistically significant difference. However, a meta-analysis comparing beta carotene and placebo groups regarding malignant transformation as a primary outcome failed to show any significant benefit. Furthermore, results showed evidence of beta carotene safety. Conclusion: the overall quality of evidence about efficacy of beta carotene in oral leukoplakia treatment was not high. However, given the obvious safety of this agent, data suggests it could have a promising effect in clinical improvement of oral leukoplakia lesions. However, no evidence supporting its benefits in reducing risk of malignant transformation in these lesions was found. Therefore, further long term, well designed randomized clinical trials are highly recommended.

Objetivos: Se realizó una revisión sistemática para evaluar la efectividad y la seguridad de los betacarotenos para el tratamiento de la leucoplasia oral en relación con la resolución clínica y la prevención de la transformación maligna. Material y Métodos: la búsqueda sistemática se realizó en tres bases de datos electrónicas y la selección del estudio se realizó de acuerdo con los criterios de elegibilidad preestablecidos. En la revisión se incluyeron cuatro estudios que evaluaban la eficacia de los betacarotenos en la leucoplasia oral en comparación con el placebo; tres de los cuales fueron asignados para el análisis cuantitativo. Los datos fueron extraídos, tabulados, su calidad evaluada y analizados estadísticamente. Resultados: El metanálisis reveló que al comparar la resolución clínica, el grupo de betacaroteno fue favorecido en comparación con el placebo, con una diferencia estadísticamente significativa. Sin embargo, un metaanálisis que comparó los grupos de betacaroteno y placebo con respecto a la transformación maligna como resultado primario no mostró ningún beneficio significativo. Además, los resultados mostraron evidencia de seguridad de betacaroteno. Conclusión: La calidad general de la evidencia sobre la eficacia del betacaroteno en el tratamiento de la leucoplasia oral no es alta. Sin embargo, dada la obvia seguridad de este agente, los datos sugieren que podría tener un efecto prometedor en la mejora clínica de las lesiones de leucoplasia oral. Sin embargo, no se encontraron pruebas que respalden sus beneficios en la reducción del riesgo de transformación maligna en estas lesiones. Por lo tanto, se recomiendan ensayos clínicos aleatorios bien diseñados a largo plazo.

Growth hormone in the tumor microenvironment

ABSTRACT Tumor development is a multistep process whereby local mechanisms enable somatic mutations during preneoplastic stages. Once a tumor develops, it becomes a complex organ composed of multiple cell types. Interactions between malignant and non-transformed cells and tissues create a tumor microenvironment (TME) comprising epithelial cancer cells, cancer stem cells, non-tumorous cells, stromal cells, immune-inflammatory cells, blood and lymphatic vascular network, and extracellular matrix. We review reports and present a hypothesis that postulates the involvement of growth hormone (GH) in field cancerization. We discuss GH contribution to TME, promoting epithelial-to-mesenchymal transition, accumulation of unrepaired DNA damage, tumor vascularity, and resistance to therapy. Arch Endocrinol Metab. 2019;63(6):568-75

Clinicopathologic characteristics of ovarian clear cell carcinoma in the background of endometrioma: a surveillance strategy for an early detection of malignant transformation in patients with asymptomatic endometrioma

OBJECTIVE: This study aimed to analyze the clinical features of clear cell carcinoma in relation to endometriosis and to determine an appropriate surveillance strategy for the early detection of malignant transformation of endometrioma in asymptomatic patients. METHODS: We retrospectively reviewed the clinicopathologic data of 50 patients with ovarian clear cell carcinoma. Clinicopathologic characteristics, treatment outcomes, and the association between endometriosis and the risk of malignant transformation were analyzed. RESULTS: Ten (20%) patients had been diagnosed with endometrioma before the diagnosis of clear cell carcinoma. The median period from the diagnosis of endometrioma to clear cell carcinoma diagnosis was 50 months (range, 12–213 months). After complete staging surgery, histological confirmation of endometriosis was possible in 35 (70%) patients. Of the 50 patients, 39 (78%) had not undergone any gynecologic surveillance until the onset of symptoms, at which time many of them presented with a rapidly growing pelvic mass (median 10 cm, range 4.6–25 cm). With the exception of 2 patients, all cancer diagnoses were made when the patients were in their late thirties, and median tumor size was found to increase along with age. Asymptomatic patients (n=11) who had regular gynecologic examinations were found to have a relatively smaller tumor size, lesser extent of tumor spread, and lower recurrence rate (P=0.011, 0.283, and 0.064, respectively). The presence of endometriosis was not related to the prognosis. CONCLUSION: Considering the duration of malignant transformation and the timing of cancer diagnosis, active surveillance might be considered from the age of the mid-thirties, with at least a 1-year interval, in patients with asymptomatic endometrioma.

Biological function and regulatory mechanism of epithelial-mesenchymal transition-related long non-coding RNAs in liver cancer / 临床肝胆病杂志

Liver cancer has a high degree of malignancy, and many oncogenes and tumor suppressor genes play an important regulatory role in the development and progression of liver cancer. Epithelial-mesenchymal transition (EMT) is a biological process in which epithelial cells transform into mesenchymal cells, which can increase the migration and invasion abilities of tumor cells. Long non-coding RNAs (lncRNAs) can regulate EMT process in various ways. This article reviews the research advances in the main biological functions and regulatory mechanisms of EMT-related lncRNAs in liver cancer.

Biological function and regulatory mechanism of epithelial-mesenchymal transition-related long non-coding RNAs in liver cancer / 临床肝胆病杂志

Liver cancer has a high degree of malignancy, and many oncogenes and tumor suppressor genes play an important regulatory role in the development and progression of liver cancer. Epithelial-mesenchymal transition (EMT) is a biological process in which epithelial cells transform into mesenchymal cells, which can increase the migration and invasion abilities of tumor cells. Long non-coding RNAs (lncRNAs) can regulate EMT process in various ways. This article reviews the research advances in the main biological functions and regulatory mechanisms of EMT-related lncRNAs in liver cancer.

A Case of Concurrent Non-Intestinal Type Adenocarcinoma and Inverted Papilloma in a Maxillary Sinus / 대한이비인후과학회지

Inverted papilloma is a benign epithelial tumor that arises from the sinonasal epithelium and occurs in 0.5–4% of all sinonasal tumors. Although benign, it is associated with malignant transformation in 2–27% of the cases, with the most commonly accompanying malignant tumor being squamous cell carcinoma. The malignant transformation of inverted papilloma into adenocarcinoma is extremely rare, with two cases reported worldwide to date. Here, along with a literature review, we report a recent case of a 53-year-old man with non-intestinal type adenocarcinoma associated with a sinonasal inverted papilloma. This case shows the possibility of a malignant transformation of inverted papilloma into non-intestinal type adenocarcinoma, which may be associated with human papilloma virus and thus requires further investigation.

Malignant transformation of abdominal wall endometriosis to clear cell carcinoma: case report

ABSTRACT BACKGROUND: Malignant transformation of endometriosis in the abdominal wall is a rare and still poorly understood event. Less than 30 cases have been reported in the worldwide literature. Most cases of solid tumors are report in a previous abdominal scar with malignant transformation of a focus of endometriosis. Presence of lymph node metastases in nearby chains is frequent and is associated with poor prognosis. CASE REPORT: We report a case of a 42-year-old woman with a history of abdominal surgery (Pfannenstiel) to resect abdominal wall endometriosis. Physical examination revealed a solid mass of approximately 10 cm x 6 cm in the anterior wall of the abdomen. Computed tomography (CT) of the abdomen and pelvis showed a heterogeneous, predominantly hypoattenuating expansive formation measuring 10.6 cm x 4.7 cm x 8.3 cm. The patient underwent exploratory incisional laparotomy, block resection of the abdominal mass and lymphadenectomy of the external and inguinal iliac chains. The abdominal wall was reconstructed using a semi-absorbable tissue-separating screen to reconstitute the defect caused by resection of the tumor. Histological evaluation revealed infiltration by malignant epithelioid neoplasia, thus confirming the immunohistochemical profile of adenocarcinoma with clear cell components. Lymphadenectomy showed metastatic involvement of an external iliac chain lymph node. CONCLUSION: Resection of the mass along with the abdominal wall, with wall margins, is the most effective treatment. Reconstruction is a challenge for surgeons. The patient has been followed up postoperatively for eight months, without any evidence of disease to date.

Expresiones clínicas de los desórdenes potencialmente malignos en cavidad oral. Revisión integrativa de la literatura / Clinical expressions of potentially malignant disorders in oral cavity. integrative review of literature

Antecedentes: Los desórdenes potencialmente malignos (DPM) son aquellas situaciones clínicas en la cavidad bucal que presentan un riesgo aumentado de malignización neoplásica, debido a la exposición a factores de riesgo o alteraciones genéticas. Es necesario realizar revisiones de la evidencia de este tipo de desórdenes para desarrollar o actualizar guías de práctica clínica idóneas. Objetivo: Identificar, a través de una revisión integrativa de la literatura, la evidencia reciente sobre DPM de la cavidad bucal y su transformación maligna, con el fin de proporcionar recomendaciones de manejo diagnóstico y terapéutico. Métodos: Se realizó una búsqueda de literatura en las bases de datos PubMed, Elsevier, SciELO y EMBASE, utilizando la combinación de seis descriptores. Resultados: La búsqueda inicial arrojó 1743 títulos y la muestra consistió en 67 artículos después de aplicar los criterios de inclusión y exclusión. Las DPM identificadas fueron liquen plano oral, palatitis nicotínica, hábito de fumar invertido, queilitis actínica, eritroplasia y leucoplasia oral y úlcera traumática crónica. Conclusión: Cada tipo de lesión tiene distinto potencial de malignización, entre los cuales la eritroplasia, el liquen plano oral variante erosivo y la queilitis actínica poseen el mayor riesgo.

Background: Potentially malignant disorders (PMD) are clinical oral cavity conditions that pose an increased risk of neoplastic malignization due to exposure to risk factors or genetic alterations. It is necessary to conduct evidence-based reviews of this type of disorders to develop or update adequate clinical practice guidelines. Purpose: Identify, through an integrative review of literature, recent evidence on PMDs in the oral cavity and their malignant transformation, in order to provide diagnostic and treatment recommendations. Methods: A literature search was carried out in the PubMed, Elsevier, SciELO, and EMBASE, using a combination of six descriptors. Results: The initial search showed 1743 titles and the sample, after applying the inclusion and exclusion criteria, consisted of 67 articles. The PMDs identified were oral lichen planus, nicotinic palatitis, inverted smoking habit, actinic cheilitis, oral erythroplakia and leukoplakia, and chronic traumatic ulcer.

Clinical characteristics and survival analysis of de novo grade 3 or transformed follicular lymphoma patients / 中华血液学杂志

Objective@#To evaluate the clinical characteristics and survival outcomes of patients with de novo grade 3 or transformed follicular lymphoma (FL).@*Methods@#Fifty-two patients treated at Peking University Cancer Hospital between January 2009 and September 2017 were assessed, including 28 patients with FL 3A grade, 13 patients with FL 3B grade, 11 patients with transformed FL. Baseline characteristics, survival and prognostic factors were analyzed.@*Results@#① Twenty-six male and 26 female patients were enrolled, including 28 patients with FL 3A grade, 13 patients with FL 3B grade, 11 patients with transformed FL. ②The 3-year progression-free survival (PFS) and overall survival (OS) for the entire cohort were 56.0% and 80.6%, respectively. Patients with international prognostic index (IPI) score 0-1 demonstrated significantly better 3-year PFS (80.3% vs 20.1%; t=18.902, P<0.001) and OS (95.7% vs 57.0%; t=10.406, P<0.001) than patients with IPI score 2-3. Three-year PFS (94.1% vs 37.2% vs 25.2%; P=0.002) and OS (100.0% vs 76.0% vs 59.8%; P=0.020) were also significantly different among patients with FLIPI 1 score 0-1, 2, ≥3. FLIPI 2 score was also identified as a prognostic factor for 3-year PFS (68.4%, 0, 0; P=0.001) and OS(87.5%, 76.2%, 0; P=0.003). ③Multivariate analysis indicated a significant association of PFS (HR=3.536, P=0.015) and OS (HR=15.713, P=0.015) with IPI. FLIPI 2 was associated with OS (score 0-1, HR=0.078, P=0.007; score 2, HR=0.080, P=0.022).@*Conclusion@#De novo grade 3 or transformed FL might be a group of curable disease with current treatment strategies. IPI is still a prognostic tool in this scenario.

Integrative analysis of gene expression profile and DNA methylation profile of long-term cultivated porcine bone marrow mesenchymal stem cells / 中国修复重建外科杂志

Objective: To integrate the result of whole genome expression data and whole genome promoter CpG island methylation data, to screen the epigenetic modulated differentially expressed genes from transformed porcine bone marrow mesenchymal stem cells (BMSCs) after long-term cultivation. Methods: Bone marrow from 6 landrace pigs, 3-month-old about 50 kg weight, was aspirated from the medullary cavity of the proximal tibia. The BMSCs were isolated, and purified by Ficoll density gradient centrifugation combined with adherent culture method. The transfor mation of BMSCs was tested by several methods including cell morphology observation, karyotype analysis, clone forming in soft agarose, serum requirement assay, and tumor forming in mice. The Agilent Pig 4x44k Gene Expression Microarray was used to investigate the differentially expressed mRNA. The methylated genes expression profile was performed using customized pig methylation chip. The gene expression and DNA methylation profiles were integrated to find out the epigenetic modulated differentially expressed genes, and to complete the bioinformatic analysis. Results: BMSCs showed a change in appearance, from the initial spindle shape to a more flatted morphology then to small contact shape. After additional passages, BMSCs gradually acquired recovery of proliferating capacity and transformation properties such as anchorage-independent growth, chromosomal abnormality, and tumor formation in nude mice. The gene chip analysis demonstrated that 257 genes were upregulated and 315 genes were downregulated during long-term cultures as well as multiple signal pathways transduction involved, such as cell cycle, ECM-receptor interaction, focal adhesion, regulation of actin cytoskeleton, pathways in cancer, and P53. The analysis from methylation chip of coding genes suggested epigenetic regulation was involved in BMSCs spontaneous transformation and play a important role on it; 962 genes were hypermethylated and 1219 genes were hypomethylated, which were involved in the biological process of cellular metabolic, structure, and tumor generation. The combined analysis of genes regulated by methylation in the transformation process of BMSCs found that the methylation changes of the 35 genes were contrary to the direction of expression change (correlation coefficient r=-0.686, P=0.000); in which the methylation level of 21 genes promoter regions were increased while the gene expression decreased, and the methylation level of the 14 genes promoter regions decreased and the gene expression increased. At the same time, KEGG enrichment analysis revealed multiple genes regulated by methylation, involved in stem cell differentiation and multiple cell signaling pathways. Among the 14 down-regulated genes, many of them have the role of regulating the interaction of tumor and immunization, and the change of the methylation status of the CDKN3 promoter region may be closely related to the cell oncology. Conclusion: The results deepen our understanding of the crucial role of coding genes methylation modification in BMSCs transformation, and may provide new approach to establish safe criteria for BMSCs clinical applications and transformation prevention.

Malignant transformation in chronic osteomyelitis / Transformação maligna na osteomielite crônica

ABSTRACT INTRODUCTION: Carcinomatous degeneration is a rare and late complication developing decades after the diagnosis of chronic osteomyelitis. OBJECTIVES: To present the results from a retrospective study of six cases of squamous cell carcinoma arising from chronic osteomyelitis. METHODS: Six cases of chronic osteomyelitis related to cutaneous squamous cell carcinoma were identified. The cause and characteristics of the osteomyelitis were analyzed, as well as time up to malignancy, the suspicion signs for malignancy, the localization and histological type of the cancer, and the type and result of the treatment. RESULTS: The mean time between osteomyelitis onset and the diagnosis of malignant degeneration was 49.17 years (range: 32-65). The carcinoma resulted from tibia osteomyelitis in five cases and from femur osteomyelitis in one. The pathological examination indicated cutaneous squamous cell carcinoma in all cases. All the patients were staged as N0M0, except for one, whose lomboaortic lymph nodes were affected. The treatment consisted of amputation proximal to the tumor in all patients. No patient presented signs of local recurrence and only one had carcinoma metastasis. CONCLUSION: Early diagnosis and proximal amputation are essential for prognosis and final results in carcinomatous degeneration secondary to chronic osteomyelitis.

RESUMO INTRODUÇÃO: Degeneração carcinomatosa é uma complicação rara e tardia que se desenvolve décadas após o diagnóstico de osteomielite crônica. OBJETIVOS: Apresentar os resultados de um estudo retrospectivo de seis casos de carcinoma espino-celular em um contexto de osteomielite crônica. MÉTODOS: Identificamos seis casos de carcinoma espino-celular relacionados à osteomielite crônica. A causa e as características da osteomielite foram analisadas, bem como o tempo decorrido até transformação maligna, os sinais de suspeita de malignização, a localização e o tipo histológico do câncer e o tipo e os resultados do tratamento. RESULTADOS: O tempo médio entre a causa da osteomielite e o diagnóstico da transformação maligna foi de 49,17 anos (intervalo: 32 a 65). O câncer teve origem em osteomielites da tíbia em cinco casos e em uma osteomielite do fêmur em um caso. A análise histológica demonstrou carcinoma espinocelular cutâneo em todos os casos. Todos os pacientes foram estadiados como N0M0, com exceção de um que apresentava atingimento dos gânglios linfáticos lomboaórticos. O tratamento foi a amputação proximal ao tumor em todos os pacientes. Nenhum dos pacientes apresentou sinais de recidiva local e apenas um desenvolveu metastização do carcinoma espinocelular. CONCLUSÃO: O diagnóstico precoce e a amputação proximal ao tumor são fundamentais para o prognóstico e os resultados finais na transformação maligna secundária a osteomielite crônica.

Espiroadenocarcinoma localizado en pabellón auricular y cuero cabelludo: reporte de un caso de localización inusual / Spiroadenocarcinoma located on ear auricle and scalp: a case report of unusual location

Introducción: El espiradenocarcinoma es una neoplasia maligna inusual que suele surgir de un espiradenoma benigno solitario preexistente. La mayoría de las lesiones aparecen en tronco y extremidades, pero casos extremadamente raros se han reportado en la región del cuero cabelludo y pabellón auricular. Objetivo: Describir el caso de un paciente en quien se diagnosticó espiradenocarcinoma. Diseño: Reporte de caso. Materiales y métodos: Se presenta el caso de un paciente adulto mayor con masa en región auricular y cuero cabelludo, de crecimiento progresivo. Resultados: Los estudios imagenológicos e histopatológicos mostraron una lesión tumoral maligna derivada de los anexos cutáneos complicada con infección y miasis. Se le informa la importancia de resección quirúrgica, pero los familiares se negaron a dicho procedimiento. Por lo cual se le ofrecen medidas paliativas. Conclusión: Describimos un caso extremadamente raro de un espiroadenocarcinoma en cuero cabelludo cerca del pabellón auricular; siendo el primer caso descrito en Colombia.

Introduction: Spiradenocarcinomas an extremely rare malignant neoplasm. Most often arises from a preexisting solitary benign spiroadenoma. Most of the lesions often appear on the trunk, limbs and unusually, on the region of the scalp near the pinna. Objective: To describe a case of a patient who was diagnosed with spiroadenoma. Design: Case report. Methods: We present the case of an elderly patient with a progressive growth mass on the scalp near the pinna. Results: Imaging studies in conjunction with histopathology allowed to evidence a malignant tumor lesion derived from skin annexes and complicated with an infection process and secondary myiasis. We told him the importance of performing surgery but the family refused this procedure. Conclusion: We report an extremely rare case of a spiroadenocarcinoma of the scalp near the pinna; this is the first case reported in Colombia