Esophagocolonic OrVil Anastomosis After Minimally Invasive Esophagectomy.

Purpose: The classical colon substitution procedure is open surgery. Still, technological developments could allow a minimally invasive procedure that might improve patient outcomes. To present the efficacy and safety of esophagocolonic OrVil anastomosis after minimally invasive esophagectomy. Methods: This retrospective study included 10 patients with esophageal cancer treated with OrVil anastomosis (OA) between August 2017 and May 2021 at Department of Thoracic Surgery, Nanjing Lishui People's Hospital, Zhongda Hospital Lishui Branch, Southeast University, Nanjing, China and the Fourth Associated Hospital of Anhui Medical University. The patient's characteristic information and related perioperative indexes were collected from the hospital's electronic medical record system and the patients were followed up. Results: The mean operative time and median intraoperative blood loss were 530 ± 88 minutes and 200 (range: 100-300) mL, respectively. A median of 26 (range: 13-30) lymph nodes was dissected per patient. The median total duration of hospitalization and postoperative hospitalization was 32 (range: 24-64) and 15 (range: 12-42) days, respectively. Seven (70%) patients had postoperative pulmonary infections. Two (20%) patients had postoperative respiratory failure. No esophagocolonic anastomotic leakage was observed in all cases. One patient was complicated with postoperative colonicoduodenal anastomotic leakage after the operation and was cured. However, 1 (10%) of the remaining 9 patients died from colonicolonic anastomotic leakage during hospitalization. The living 9 cases were followed up, and the median overall survival time was 36 months. Conclusion: Colonic interposition for esophageal cancer is effective and safe using the minimally invasive OA technique.

DAB2IP predicts treatment response and prognosis of ESCC patients and modulates its radiosensitivity through enhancing IR-induced activation of the ASK1-JNK pathway.

BACKGROUND: Disabled homolog 2 interacting protein (DAB2IP) plays a tumor-suppressive role in several types of human cancers. However, the molecular status and function of the DAB2IP gene in esophageal squamous cell carcinoma (ESCC) patients who received definitive chemoradiotherapy is rarely reported. METHODS: We examined the expression dynamics of DAB2IP by immunohistochemistry (IHC) in 140 ESCC patients treated with definitive chemoradiotherapy. A series of in vivo and in vitro experiments were performed to elucidate the effect of DAB2IP on the chemoradiotherapy (CRT) response and its underlying mechanisms in ESCC. RESULTS: Decreased expression of DAB2IP in ESCCs correlated positively with ESCC resistance to CRT and was a strong and independent predictor for short disease-specific survival (DSS) of ESCC patients. Furthermore, the therapeutic sensitivity of CRT was substantially increased by ectopic overexpression of DAB2IP in ESCC cells. In addition, knockdown of DAB2IP dramatically enhanced resistance to CRT in ESCC. Finally, we demonstrated that DAB2IP regulates ESCC cell radiosensitivity through enhancing ionizing radiation (IR)-induced activation of the ASK1-JNK signaling pathway. CONCLUSIONS: Our data highlight the molecular etiology and clinical significance of DAB2IP in ESCC, which may represent a new therapeutic strategy to improve therapy and survival for ESCC patients.

Long non-coding RNA LINC01270 is an onco-promotor in lung adenocarcinoma by upregulating LARP1 via sponging miR-326.

Accumulating evidence have proved the key role of long non-coding RNA in lung adenocarcinoma (LUAD) progression. Bioinformatics analysis is used to seek the differentially expressed lncRNA LINC01270 from TCGA database. The overexpression of LINC01270 was then verified in LUAD tumor tissues and cell lines by qRT-PCR. LINC01270 knockdown resulted in impaired cell proliferative and invasive ability via CCK-8 assay, EdU assay, colony formation assay, transwell assay, while aberrant upregulation of LINC01270 led to enhanced cell growth and invasion. Moreover, LINC01270 was found inhibiting miR-326 and thereby overexpressing the abundance of LARP1 to promote LUAD development via PI3K/AKT pathway. It was also proved that LINC01270 knockdown could suppress LUAD tumor growth in vivo. All of these findings demonstrate thatLINC01270 is a tumor promotor in LUAD via enhancing LARP1 expressed by sponging miR-326 to facilitate the development of LUAD. LINC01270 play a significant role in LUAD, which could serve as biomarkers for early diagnosis and a novel targeted remedy.

The comparisons of three stapler placement methods for intrathoracic mechanistic circular stapling in Ivor Lewis minimally invasive esophagectomy.

BACKGROUND: To analyze the impact of the reversal penetrating technique (RPT) for intrathoracic gastroesophageal mechanical anastomosis on the development of anastomotic complications in Ivor Lewis minimally invasive esophagectomy (ILMIE), and to further identify the risk factors for the development of anastomotic leakage and stricture. METHODS: A retrospective observational study was conducted using the clinical data of 316 patients with esophageal carcinoma (EC) who underwent ILMIE from January 2012 to December 2019. The participants were divided into three groups, namely the RPT group, the transoral Orvil technique (TOT) group, and the purse-string technique (PST) group, according to the different stapler placement methods for intrathoracic mechanistic circular stapling. Multivariate analysis was performed to investigate the association of risk factors with anastomotic leakage and stricture. RESULTS: There were 154 patients in the RPT group, 78 in the TOT group, and 84 in the PST group for intrathoracic gastroesophageal circular stapling in ILMIE. There were no differences in intraoperative anastomosis-related conditions including conversion of open operations, and lymph nodes harvested between the three groups. However, the mean total operative time and gastroesophageal anastomosis time in the RPT group were significantly shorter than those in the other groups (both P<0.05). The rates of anastomotic leakage and stricture showed no statistical differences between the three groups (leakage: P=0.875; stricture: P=0.942). Multivariate analysis revealed that the RPT method of anvil placement did not increase the probability of anastomotic leakage [RPT: reference; TOT: odds ratio (OR) 0.422, P=0.341; PST: OR 1.436, P=0.645] and stricture (RPT: reference; TOT: OR 0.579, P=0.376; PST: OR 1.195, P=0.755). CONCLUSIONS: The RPT method of anvil placement for intrathoracic gastroesophageal circular stapling does not increase the risk of anastomotic complications in ILMIE, but had significantly shorter surgical time and anastomosis time.

Circ-HACE1 Aggravates Cigarette Smoke Extract-Induced Injury in Human Bronchial Epithelial Cells via Regulating Toll-Like Receptor 4 by Sponging miR-485-3p.

BACKGROUND: Smoking is the most common cause of chronic obstructive pulmonary disease (COPD), and the early diagnosis for COPD remains poor. Exploring the molecular mechanism and finding feasible biomarkers will be beneficial for clinical management of COPD. Circular RNAs (circRNAs) are noncoding RNAs that act as miRNA sponges to regulate the expression levels of genes, leading to the changes of cellular phenotypes and disease progression. CircRNA HECT domain and ankyrin repeat containing E3 ubiquitin protein ligase 1 (circ-HACE1) was abnormally expressed after the induction of cigarette smoke extract (CSE) in cell model. This study was performed to explore its function and mechanism in COPD. METHODS: Circ-HACE1, microRNA-485-3p (miR-485-3p) and toll-like receptor 4 (TLR4) detection was performed by quantitative real-time polymerase chain reaction (qRT-PCR). Cell viability and apoptosis/cell cycle were respectively examined using cell counting kit-8 (CCK-8) and flow cytometry. Inflammatory cytokines were determined by enzyme-linked immunosorbent assay (ELISA). Oxidative stress was evaluated through the measurement of malondialdehyde (MDA) and superoxide dismutase (SOD). The target binding analysis was conducted via dual-luciferase reporter assay. Western blot was employed for protein expression detection of TLR4. RESULTS: Circ-HACE1 was overexpressed in smokers or smokers with COPD and CSE upregulated circ-HACE1 expression in 16HBE cells. Knockdown of circ-HACE1 attenuated CSE-stimulated cell viability and cell cycle repression, as well as the enhancement of cell apoptosis, inflammatory response and oxidative stress. MiR-485-3p was a target of circ-HACE1. Circ-HACE1 regulated CSE-induced cell injury via targeting miR-485-3p. TLR4 was a downstream target of miR-485-3p, and miR-485-3p inhibited the CSE-induced cell damages by directly downregulating the level of TLR4. Circ-HACE1/miR-485-3p regulated TLR4 expression in CSE-treated 16HBE cells, and TLR4 overexpression also reversed all effects of si-circ-HACE1 on CSE-treated 16HBE cells. CONCLUSION: These findings elucidated that circ-HACE1 contributed to the CSE-induced cell damages in COPD cell models via regulating TLR4 by acting as the sponge of miR-485-3p.

Comparison of Two Circular-Stapled Techniques for Esophageal Cancer: A Propensity-Matched Analysis.

OBJECTIVE: The optimal technique for the thoracoscopic construction of an intrathoracic esophagogastric anastomosis continues to be a subject of controversy. The aim of this study was to compare the perioperative outcomes of circular-stapled anastomosis using a transorally inserted anvil (Orvil™) with those of circular-stapled anastomosis using a transthoracically placed anvil (non-Orvil™) in totally minimally invasive Ivor Lewis esophagectomy (Ivor Lewis TMIE). METHODS: The data of 272 patients who underwent Ivor Lewis TMIE for esophageal cancer at multiple centers were collected from January 1, 2014 to December 31, 2017. After propensity score matching (1:1) for patient baseline characteristics, 65 paired cases were selected for statistical analysis. Logistic regression analysis was performed to investigate the significant factors of anastomotic leakage. RESULTS: In the propensity score-matched analysis, compared with the non-Orvil™ group, the Orvil™ group was associated with a significantly shorter operation time (p=0.031), less intraoperative hemorrhage (p<0.001), lower need for intraoperative transfusions (p=0.009), earlier postoperative oral feeding time (p=0.010), longer chest tube duration (p<0.001), shorter postoperative hospital stays (p=0.001), lower total hospitalization costs (p<0.001) and a lower postoperative anastomotic leakage rate (p=0.033). Multivariate logistic regression analysis showed that anastomotic technique and pulmonary infection were independent factors for the development of postoperative anastomotic leakage (p< 0.05). CONCLUSIONS: Orvil™ anastomosis exhibited better perioperative effects than non-Orvil™ anastomosis after the propensity score-matched analysis. Remarkably, the Orvil™ technique contributed to a lower postoperative anastomotic leakage rate than the non-Orvil™ technique.

Identification of the tropomyosin (HL-Tm) in Haemaphysalis longicornis.

Haemaphysalis longicornis tropomyosin (HL-Tm) was amplified by RT-PCR. The cDNA contained a 825 bp open reading frame coding for 274 amino acids with a predicted theoretical isoelectric point (pI) of 4.55 and molecular weight of 31.7 kDa. Real-time RT-PCR analysis showed that the expression levels of the HL-Tm in the unfed-females were significantly higher than in other tested developmental stages (eggs, unfed-larvae and unfed-nymphs). Western blot analysis showed that rabbit anti-serum against H. longicornis unfed-adult ticks recognized the recombinant HL-Tm protein (rHL-Tm). Immunization of rabbits with the rHL-Tm resulted in a statistically significant reduction of female engorgement and oviposition. Silencing of HL-Tm by RNAi showed a decrease in tick engorgement and oviposition, which is consistent with the effect of recombinant protein vaccine on the adults. These results showed that tick HL-Tm might be involved in the regulation of ticks blood-feeding, growth and oviposition.

Hybrid video-assisted thoracic surgery with segmental-main bronchial sleeve resection for non-small cell lung cancer.

BACKGROUND: The purpose of the current study is to present the clinical and surgical results in patients who underwent hybrid video-assisted thoracic surgery with segmental-main bronchial sleeve resection. METHODS: Thirty-one patients, 27 men and 4 women, underwent segmental-main bronchial sleeve anastomoses for non-small cell lung cancer between May 2004 and May 2011. RESULTS: Twenty-six (83.9%) patients had squamous cell carcinoma, and 5 patients had adenocarcinoma. Six patients were at stage IIB, 24 patients at stage IIIA, and 1 patient at stage IIIB. Secondary sleeve anastomosis was performed in 18 patients, and Y-shaped multiple sleeve anastomosis was performed in 8 patients. Single segmental bronchiole anastomosis was performed in 5 cases. The average time for chest tube removal was 5.6 days. The average length of hospital stay was 11.8 days. No anastomosis fistula developed in any of the patients. The 1-, 2-, and 3-year survival rates were 83.9%, 71.0%, and 41.9%, respectively. CONCLUSION: Hybrid video-assisted thoracic surgery with segmental-main bronchial sleeve resection is a complex technique that requires training and experience, but it is an effective and safe operation for selected patients.

PITX2: a promising predictive biomarker of patients' prognosis and chemoradioresistance in esophageal squamous cell carcinoma.

The paired-like homeodomain transcription factor 2 (PITX2), a downstream effector of wnt/ß-catenin signaling, is well known to play critical role during normal embryonic development. However, the possible involvement of PITX2 in human tumorigenesis remains unclear. In this study, we extend its function in human esophageal squamous cell carcinoma (ESCC). The real-time PCR, Western blotting and immunohistochemistry (IHC) methods were applied to examine expression pattern of PITX2 in two different cohorts of ESCC cases treated with definitive chemoradiotherapy (CRT). Receiver operating characteristic (ROC) curve analysis was used to determine the cutoff point for PITX2 high expression in the training cohort. The ROC-derived cutoff point was then subjected to analyze the association of PITX2 expression with patients' survival and clinical characteristics in training and validation cohort, respectively. The expression level of PITX2 was significantly higher in ESCCs than that in normal esophageal mucosa. There was a positive correlation between PITX2 expression and clinical aggressiveness of ESCC. Importantly, high expression of PITX2 was observed more frequently in CRT resistant group than that in CRT effective group (p < 0.05). Furthermore, high expression of PITX2 was associated with poor disease-specific survival (p < 0.05) in ESCC. Then, the MTS, clonogenic survival fraction and cell apoptosis experiments showed that knockdown of PITX2 substantially increased ESCC cells sensitivity to ionizing radiation (IR) or cisplatin in vitro. Thus, the expression of PITX2, as detected by IHC, may be a useful tool for predicting CRT resistance and serves as an independent molecular marker for poor prognosis of ESCC patients treated with definite CRT.

HIF-1α effects on angiogenic potential in human small cell lung carcinoma.

BACKGROUND: Hypoxia-inducible factor-1 alpha (HIF-1α) maybe an important regulatory factor for angiogenesis of small cell lung cancer (SCLC). Our study aimed to investigate the effect of HIF-1α on angiogenic potential of SCLC including two points: One is the effect of HIF-1α on the angiogenesis of SCLC in vivo. The other is the regulation of angiogenic genes by HIF-1α in vitro and in vivo. METHODS: In vivo we used an alternative method to study the effect of HIF-1a on angiogenic potential of SCLC by buliding NCI-H446 cell transplantation tumor on the chick embryo chorioallantoic membrane (CAM) surface. In vitro we used microarray to screen out the angiogenic genes regulated by HIF-1a and tested their expression level in CAM transplantation tumor by RT-PCR and Western-blot analysis. RESULTS: In vivo angiogenic response surrounding the SCLC transplantation tumors in chick embryo chorioallantoic membrane (CAM) was promoted after exogenous HIF-1α transduction (p < 0.05). In vitro the changes of angiogenic genes expression induced by HIF-1α in NCI-H446 cells were analyzed by cDNA microarray experiments. HIF-1α upregulated the expression of angiogenic genes VEGF-A, TNFAIP6, PDGFC, FN1, MMP28, MMP14 to 6.76-, 6.69-, 2.26-, 2.31-, 4.39-, 2.97- fold respectively and glycolytic genes GLUT1, GLUT2 to2.98-, 3.74- fold respectively. In addition, the expression of these angiogenic factors were also upregulated by HIF-1α in the transplantion tumors in CAM as RT-PCR and Western-blot analysis indicated. CONCLUSIONS: These results indicated that HIF-1α may enhance the angiogenic potential of SCLC by regulating some angiogenic genes such as VEGF-A, MMP28 etc. Therefore, HIF-1α may be a potential target for the gene targeted therapy of SCLC.

[An indirect ELISA for the detection of Babesia caballi in equine animals].

OBJECTIVE: To clone and express BC48 gene of Babesia caballi, and to establish an indirect ELISA for the diagnosis of B. caballi in equine animals. METHOD: The genomic DNA of B. caballi was extracted from the infected donkey blood. BC48 gene was amplified by PCR. The PCR product was cloned into expression plasmid pET28a, and expressed in E. coli BL21 with IPTG induction. The recombinant protein was purified by Ni-NTA affinity chro-matography and was used as a diagnostic antigen to establish an indirect ELISA. The reaction conditions of the indirect ELISA were optimized. Specificity and sensitivity of this method were evaluated. RESULT: BC48 gene of B. caballi was 1 272 bp. The recombinant protein was expressed in E. coli BL21 as a soluble protein with a molecular weight of about M, 46 000 under induction of IPTG. The concentration of purified protein was 12.98 mg/ml. The best conditions were obtained for the ELISA when the antigen concentration was 65 microg/ml with the serum dilution of 1:80. The protein specifically reacted with serum from donkey infected by B. caballi, but did not react with serum from donkey infected by Theileria equi (B. equi). Both ELISA and microscopy were applied to examine 17 donkeys in the field, 3 were positive by ELISA and 2 were found parasite-positive, respectively. CONCLUSION: The indirect ELISA method may be used to detect B. caballi infection in equine animals.

[Construction of cDNA expression library of unfed female Haemaphysalis longicornis and immuno-screening].

OBJECTIVE: To construct a cDNA expression library from unfed female tick Haemaphysalis longicornis for screening and cloning potential antigenic genes. METHODS: Total RNA was isolated from unfed female ticks, mRNA was purified and a library of oligo (dT) -primed cDNA with added directional EcoR I /Hind III linkers was constructed from the purified mRNA. The constructed cDNA was ligated to the EcoR I /Hind III arms of the lambda SCREEN vector. Pure phage stocks were harvested by plaque purification and converted to plasmid subclones by plating phage on host strain BM25.8. Recombinant plasmids that were subcloned to E. coli BM25.8 were isolated and transformed into E. coli JM109. Recombinant plasmids abstracted from JM109 were identified by PCR and sequencing. RESULTS: The recombinant phage DNA was packaged by using phage-marker packaging extracts, resulting in a primary cDNA library with a size of 1.8 x 10(6) pfu. Data showed 100% of the library were recombinant and the titer of the amplified library was 2.4 x 10(9) pfu/ml. Forty-two clones of encoding immunodominant antigens were obtained from the cDNA library. Sequence analysis revealed 12 unique cDNA sequences and the encoded putative proteins showed similarities to H. longicornis tropomyosin mRNA, Rhipicephalus annulatus unknown larval protein mRNA, chromosome 2R of Drosophila melanogaster, mitochondrial DNA of H. flava, clones HqL09 unkown mRNA and Hq05 mRNA of H. qinghaiensis, and myosin alkali light chain protein mRNA. CONCLUSION: The cDNA expression library from unfed female H. longicornis was successfully constructed and screening of protective genes may provide candidate antigens of the tick.