Lactoferrin and its digestive peptides induce interferon-α production and activate plasmacytoid dendritic cells ex vivo.

Plasmacytoid dendritic cells (pDCs) recognise viral single-stranded RNA (ssRNA) or CpG DNA via Toll-like receptor (TLR)-7 and TLR9, and produce interferon (IFN)-α. Activated pDCs upregulate human leukocyte antigen (HLA)-DR and CD86 expression levels. Ingestion of bovine lactoferrin (LF) activates pDCs, but little is known about its effects. In this study, the effects of LF and its pepsin hydrolysate (LFH) on the production of IFN-α from peripheral blood mononuclear cells (PBMCs) and pDCs were examined. PBMCs were prepared from peripheral blood of healthy adults and incubated with LF, LFH, or lactoferricin (LFcin) in the absence or presence of ssRNA derived from human immunodeficiency virus. The concentration of IFN-α in the supernatant and the expression levels of IFN-α, HLA-DR, and CD86 in pDCs were quantified by enzyme-linked immunosorbent assay and flow cytometry. In the absence of ssRNA, the concentration of IFN-α was negligible and LF had no effect on it. In the presence of ssRNA, IFN-α was detected at a certain level, and LF and LFH significantly increased its concentration. The increase caused by LFH and LFcin were comparable. In addition, LF significantly upregulated the expression levels of IFN-α, HLA-DR, and CD86 in pDCs. LF and its digestive peptides induced IFN-α production and activated pDCs in the presence of ssRNA, suggesting that LF modulates the immune system by promoting pDC activation upon viral recognition.

Circulating Tumor DNA as a Potential Prognostic Marker in Patients with Borderline-Resectable Pancreatic Cancer Undergoing Neoadjuvant Chemotherapy Followed by Pancreatectomy.

BACKGROUND: Circulating tumor DNA (ctDNA) might be a promising biomarker for pancreatic cancer in liquid biopsy. This study aimed to evaluate the usefulness of liquid biopsy for patients with borderline-resectable pancreatic cancer (BR-PC). METHODS: Patients with BR-PC according to the National Comprehensive Cancer Network guidelines (2017) and eligible for neoadjuvant chemotherapy (NAC) followed by pancreatectomy were recruited at Wakayama Medical University Hospital (UMIN000026647) between March 2017 and April 2020. The study enrolled 55 patients with locally advanced PC, and each patient consented to inclusion in the study. The study investigated the relationship between KRAS status in ctDNA and clinicopathologic features, analyzing ctDNA at three time points: pretreatment, post-NAC, and post-operation. RESULTS: Of the 55 enrolled patients with a diagnosis of BR-PC, 34 were scheduled to undergo pancreatectomy. From 27 patients with resected BR-PC, 81 blood samples were analyzed in triplicate for ctDNA. The patients with positive pretreatment and post-NAC ctDNA status had no significant decrease in median relapse-free survival (RFS) or overall survival (OS). However, the patients with positive postoperation ctDNA status had a significantly shorter median OS (723 days) than the patients with negative ctDNA results (not reached; P = 0.0148). A combined analysis of postoperative ctDNA and CA19-9 values showed the cumulative effect on both RFS (P = 0.0066) and OS (P = 0.0046). The adjusted hazard ratio for risk of survival computed for the patients carrying risk factors (either detectable ctDNA or CA19-9 > 37 U/ml) increased from 4.13-fold to 17.71-fold (both P = 0.0055) compared with the patients who had no risk factors. CONCLUSION: Positive ctDNA predicts poor survival for patients with BR-PC who undergo NAC followed by pancreatectomy.

Immunomodulatory activity of heat-killed Lacticaseibacillus paracaseiMCC1849 based on the activation of plasmacytoid dendritic cells in the peripheral blood of healthy adults.

Probiotics are widely used in food for their health benefits to the host. Inactivated probiotics also reportedly improve the intestinal environment and immune regulation. Our previous studies showed that heat-killed Lacticaseibacillus paracasei MCC1849 (hk-MCC1849) effectively induced IL-12 production in mouse spleen cells and significantly reduced cold symptoms in clinical trial subjects. To further elucidate the mechanism of host immune regulation by hk-MCC1849, human peripheral blood mononuclear cells (PBMCs) were cocultured with hk-MCC1849. The Toll-like receptor 9 ligands CpG-ODN 2216 and hk-MCC1849 and the heat-killed Lacticaseibacillus rhamnosus ATCC53103 were used as positive and negative controls, respectively. The results showed that, compared with the control, hk-MCC1849 significantly increased the expression of the plasmacytoid dendritic cell (pDC) marker CD86 (p < .0001) and the pDC marker HLA-DR (p < .001) in PBMCs. The expression levels of the IL-12p40, IFNα, IFNα1, IFNγ, and ISG15 genes were significantly increased after coculture with hk-MCC1849 (p < .05, p < .05, p < .05, p < .05, and p < .05, respectively, vs. control). Furthermore, to confirm whether hk-MCC1849 directly interacted with pDCs, DCs were enriched with PBMCs following 24 h of coculture with hk-MCC1849. Phagocytosis of fluorescently labeled hk-MCC1849 by pDCs was observed, and there were significant increases in CD86 (p < .05) and HLA-DR (p < .0001) expression in pDCs. These results suggest that hk-MCC1849 exerts a potential immunomodulatory effect on the host through the activation of peripheral pDCs.

Identification of a urinary CD276 fragment for detecting resectable pancreatic cancer using a C-terminal proteomics strategy.

This study aimed to confirm urinary protein fragments in relation to the presence of pancreatic ductal adenocarcinoma (PDAC) via a C-terminal proteomics strategy using exploratory and validation cohorts. Urinary fragments were examined by iTRAQ-labelling of tryptic peptides and concentrations of C-terminal fragments were evaluated. Only the urinary CD276 fragment showed a fold change (FC) of > 1.5 with a significant difference of P < 0.01 between healthy (H) and PDAC participants in both the exploratory (H, n = 42; PDAC, n = 39) and validation cohorts (H, n = 36; resectable PDAC, n = 28). The sensitivity and specificity of the CD276 fragment for diagnosing resectable PDAC were 75% and 89%, respectively, in the validation cohort. Postoperative urinary levels of the CD276 fragment were low as compared to those before surgery (n = 18, P < 0.01). Comprehensive C-terminus proteomics identified an increase in the urinary CD276 fragment level as a feature of patients with PDAC. The urinary CD276 fragment is a potential biomarker for detecting resectable PDAC.

Scoring system to predict positive peritoneal cytology in patients with resectable and borderline resectable pancreatic cancer.

BACKGROUND: The aim of this study was to evaluate factors to predict positive peritoneal cytology, whcih would determine the indication for staging laparoscopy in pancreatic cancer. METHODS: A total of 430 patients that underwent pancreatectomy for resectable and borderline resectable pancreatic cancer were retrospectively reviewed. RESULTS: Among 430 patients, 36 had positive cytology (8.4%). Median survival time in negative cytology was 24.7 months, compared with 15.1 months in positive cytology (p = .004). Factors to predict positive cytology in pancreatic cancer according to multivariate analysis were tumor location (body, tail; OR 2.66; 95% CI: 1.21-5.85; p = .015), tumor size ≥30 mm (OR 2.95; 95% CI: 1.35-6.47; p = .007) and radiographic other-organ invasion (HR 2.79; 95% CI: 1.01-7.67; p = .047). Patients were scored 0 to 3 corresponding with these factors. Rates of positive cytology increases in each score were: score 0: 2.9%, score 1: 6.7%, score 2: 18.3%, score 3: 36.8%. CONCLUSIONS: Tumor location (body or tail), tumor size ≥30 mm, and radiographic other-organ invasions were risk factors for positive cytology in pancreatic cancer. This scoring system might be a useful indicator to perform staging laparoscopy to diagnose positive cytology.

Short-term serial circulating tumor DNA assessment predicts therapeutic efficacy for patients with advanced pancreatic cancer.

PURPOSE: We investigated the potential clinical utility of short-term serial KRAS-mutated circulating cell-free tumor DNA (ctDNA) assessment for predicting therapeutic response in patients undergoing first-line chemotherapy for advanced pancreatic cancer. METHODS: We collected 144 blood samples from 18 patients with locally advanced or metastatic cancer that were undergoing initial first-line chemotherapy of gemcitabine plus nab-paclitaxel (GEM plus nab-PTX). Analysis of KRAS-mutated ctDNA was quantified by digital droplet polymerase chain reaction (ddPCR) as mutant allele frequency (MAF). This study investigated pretreatment KRAS-mutated ctDNA status and ctDNA kinetics every few days (days 1, 3, 5 and 7) after initiation of chemotherapy and their potential as predictive indicators. RESULTS: Of the 18 enrolled patients, an increase in KRAS-mutated ctDNA MAF values from day 0-7 after initiation of chemotherapy was significantly associated with disease progression (P < 0.001). Meanwhile, positive pretreatment ctDNA status (MAF ≥ 0.02%) (P = 0.585) and carbohydrate antigen 19-9 (CA19-9) values above the median (P = 0.266) were not associated with disease progression. In univariate analysis, this short-term increase in ctDNA MAF values (day 0-7) was found to be associated with significantly shorter progression free survival (PFS) (hazard ration [HR], 24.234; range, (2.761-212.686); P = 0.0002). CONCLUSION: This short-term ctDNA kinetics assessment may provide predictive information to reflect real-time therapeutic response and lead to effective refinement of regimen in patients with advanced pancreatic cancer undergoing systemic chemotherapy.

Positive drain fluid culture on postoperative day 1 predicts clinically relevant pancreatic fistula in early drain removal with higher drain fluid amylase after pancreaticoduodenectomy.

BACKGROUND: This study aimed to clarify the risk factors of clinically relevant pancreatic fistula after early drain removal with higher drain fluid amylase after pancreaticoduodenectomy. Clinical evaluation of early drain removal with a higher drain fluid amylase after pancreaticoduodenectomy has been controversial. The safety and effectiveness have not been sufficiently examined. METHODS: Between 2015 and 2020, prophylactic surgical drains were prospectively removed on postoperative day 4 regardless of drain fluid amylase level in 364 study-eligible patients who underwent pancreaticoduodenectomy. Patients were classified according to drain fluid amylase on postoperative day 1: 281 patients with drain fluid amylase <4,000 U/L, and 83 patients with drain fluid amylase ≥4,000 U/L. RESULTS: Clinically relevant pancreatic fistula occurred in 40 of 364 enrolled patients (11.0%). In the entire cohort, male, positive postoperative day 1 drain fluid culture, and postoperative day 1 drain fluid amylase ≥4,000 U/L were independent risk factors for clinically relevant pancreatic fistula after early drain removal. When stratifying by 4,000 U/L of postoperative day 1 drain fluid amylase, the rate of clinically relevant pancreatic fistula in postoperative day 1 drain fluid amylase <4,000 U/L was significantly lower than that in postoperative day 1 drain fluid amylase ≥4,000 U/L (4% vs 35%, P < .001) after early drain removal. Moreover, in postoperative day 1 drain fluid amylase <4,000 U/L, positive postoperative day 1 drain fluid culture did not develop clinically relevant pancreatic fistula after early drain removal. However, in postoperative day 1 drain fluid amylase ≥4,000 U/L, multivariate analysis clarified that positive postoperative day 1 drain fluid culture was the only independent risk factor of clinically relevant pancreatic fistula after early drain removal (odds ratio 26.27, 95% confidence interval 5.59-123.56, P = .001). CONCLUSION: Positive drain fluid culture on postoperative day 1 might predict clinically relevant pancreatic fistula in early drain removal with a higher drain fluid amylase.

The Potential Immunomodulatory Effect of Bifidobacterium longum subsp. longum BB536 on Healthy Adults through Plasmacytoid Dendritic Cell Activation in the Peripheral Blood.

The interaction between the gut microbiota and the host can influence the host's immune system. Bifidobacterium, a commensal genus of gut bacteria, seems to have positive effects on host health. Our previous clinical research showed that B. longum subsp. longum BB536 enhanced innate and adaptive immune responses in elderly individuals with a lower grade of immunity, but the immunomodulatory mechanism is still unclear. In this study, dendritic cell (DC) surface markers in peripheral blood mononuclear cells isolated from healthy individuals were evaluated through coculture with heat-killed BB536. DC markers, innate immune activity and cytokine levels in plasma were also evaluated by a randomized, double-blind, placebo-controlled, parallel-group study (UMIN000045564) with 4 weeks of continuous live BB536 intake. BB536 significantly increased the expression of CD86 and HLA-DR on plasmacytoid DCs (pDCs) in vitro. Compared to placebo (n = 48), a significant increase in the expression of CD86 on peripheral pDCs was detected at week 4 of live BB536 intake (n = 49; 1 × 1010 CFU/day). Furthermore, coculture with hk-BB536 significantly increased the IFNγ expression level and demonstrated trends of increased IFNα1 and IFNß expression. These findings suggest that consumption of BB536 has potential immunomodulatory effects on healthy individuals through the activation of peripheral pDCs.

Radiographic duodenal invasion is associated with poor prognosis and early recurrence in patients with pancreatic ductal adenocarcinoma.

BACKGROUND: The prognostic impact of radiographic duodenal invasion (rDI) of pancreatic ductal adenocarcinoma (PDAC) has yet to be fully elucidated. This retrospective study aimed to investigate the prognostic and clinicopathological significance of rDI in patients with PDAC after pancreatoduodenectomy (PD). MATERIALS AND METHODS: We retrospectively analyzed 223 consecutive patients with resectable (R) and borderline resectable (BR)-PDAC that underwent up-front PD between 2002 and 2018. rDI was assessed by preoperative multi-detector row computed tomography. RESULTS: Ninety-three (42%) patients with PDAC had rDI, and all of them had pathological DI (pDI). The rDI(+) group had larger tumor size, BR-PDAC was more common, there was higher serum CA19-9 level, and microscopic lymphovascular invasion was more common than in the rDI(-) group. rDI was associated with significant reduction in overall survival (OS) (P < 0.001) and recurrence-free survival (RFS) (P < 0.001). In multivariate analysis, rDI was an independent prognostic factor in OS [hazard ratio (HR) = 0.52; 95% confidence interval (CI) 0.38-0.73, P < 0.001] and RFS [HR = 0.56; 95% CI 0.40-0.78, P = 0.001]. rDI was also an independent risk factor for early recurrence within 12 months [odds ratio (OR) = 0.36; 95% CI 0.18-0.73, P = 0.005]. rDI had positive correlation with liver recurrence (P = 0.024). CONCLUSION: Biological aggressiveness of PDAC with rDI implies short OS and early recurrence with frequent liver metastasis. Aggressive perioperative chemotherapy is recommended to improve prognosis, especially for R-PDAC patients with rDI.

B cell receptor repertoire analysis from autopsy samples of COVID-19 patients.

Neutralizing antibodies against the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are being developed world over. We investigated the possibility of producing artificial antibodies from the formalin fixation and paraffin-embedding (FFPE) lung lobes of a patient who died by coronavirus disease 2019 (COVID-19). The B-cell receptors repertoire in the lung tissue where SARS-CoV-2 was detected were considered to have highly sensitive virus-neutralizing activity, and artificial antibodies were produced by combining the most frequently detected heavy and light chains. Some neutralizing effects against the SARS-CoV-2 were observed, and mixing two different artificial antibodies had a higher tendency to suppress the virus. The neutralizing effects were similar to the immunoglobulin G obtained from healthy donors who had received a COVID-19 mRNA vaccine. Therefore, the use of FFPE lung tissue, which preserves the condition of direct virus sensitization, to generate artificial antibodies may be useful against future unknown infectious diseases.

Safety and feasibility of laparoscopic gastrectomy for remnant gastric cancer compared with open gastrectomy: Single-center experience.

ABSTRACT: The usefulness, safety and oncological validity of laparoscopic gastrectomy (LG) for remnant gastric cancer (RGC) have not been widely reported.A total of 38 patients who underwent gastrectomy for RGC were enrolled at Wakayama Medical University Hospital between April 2008 and December 2018. All consecutive patients were included in this retrospective study; the patients were divided into the open gastrectomy group and the laparoscopic group according to the sequential nature of their operation. Fifteen patients underwent open gastrectomy for RGC (OGR) between April 2008 and December 2013, and 23 patients underwent LG for RGC (LGR) after 2014.In the OGR group, all initial operations were performed by open surgery, whereas in the LGR group, 11 patients (47%) initially underwent laparoscopic surgery and 12 patients (53%) initially underwent open surgery (P = .002), 3 patients of which (25%) converted to open gastrectomy. There was no significant difference in the number of lymph node dissections or in operative time between the 2 groups, but blood loss was significantly lower in the LGR group than that in the OGR group (P = .002). Furthermore, although there was no difference between the 2 groups in C-reactive protein value on postoperative day 1, C-reactive protein value on postoperative day 3 was significantly lower in the LGR group than in the OGR group (P = .012). There were no differences in postoperative complications or long-term outcomes, including recurrence-free survival and overall survival.LGy is suitable in cases in which the initial surgery is performed by laparoscopic surgery. Even if the initial surgery is open surgery, it is oncologically equivalent to open gastrectomy and can be performed safely with less blood loss.

Single-incision laparoscopic antrectomy for type I gastric neuroendocrine tumor: a case report.

BACKGROUND: Type I gastric neuroendocrine tumors (GNETs) originate from hyperplasia of enterochromaffin-like (ECL) cells and are commonly detected in patients with chronic atrophic gastritis, including autoimmune gastritis. Typical treatment for type I GNETs comprises simple surveillance and/or endoscopic resection. For alleviation of hypergastrinemia resulting in ECL cell hypertrophy, antrectomy is a treatment option. Type I GNETs mostly have excellent prognosis, and if a surgical approach is chosen, the procedure must be minimally invasive. One such technique for multiple type I GNETs, minimally invasive single-incision laparoscopic antrectomy (SILA), is reported here for the first time. CASE PRESENTATION: We performed SILA on a 46-year-old woman who developed type I GNETs caused by hypergastrinemia due to autoimmune gastritis. A Lap-Protector was inserted in a 3 cm incision at the umbilicus, and set an EZ Access equipped with two 5 mm trocars and one 12 mm trocar. Antrectomy without lymph node dissection was performed using a 5 mm forward-oblique viewing endoscope, a vessel sealing device, and linear staplers, while reconstruction was by Billroth I reconstruction. Side-to-side anastomosis was performed using a 45 mm linear stapler. The stapler entry hole was sutured intracorporeally using barbed suture material. The operation time was 140 min and blood loss was 5 ml. The patient was discharged ten days after surgery without complications. Serum gastrin level decreased to within the normal range on the day after the operation. One year after surgery, esophagogastroduodenoscopy showed pathological disappearance of all lesions of the remnant stomach. CONCLUSIONS: SILA is a minimally-invasive and tolerable technique for treatment of multiple type I GNETs. In this reported case there was good cohesiveness and effectiveness in normalizing gastrin levels and in elimination of remnant gastric lesions.

Esophagogastrostomy With Fundoplication Versus Double-tract Reconstruction After Laparoscopic Proximal Gastrectomy for Gastric Cancer.

PURPOSE: We investigated that double-tract reconstruction (DTR) may be more beneficial than esophagogastrostomy (EG) with fundoplication in terms of nutritional outcomes, focusing on loss of body weight. MATERIALS AND METHODS: This study included 56 consecutive patients with early gastric cancer in the upper third of the stomach who received laparoscopic proximal gastrectomy, 39 underwent EG. In the 17 patients requiring resection of the abdominal esophagus or where the size of the remnant stomach was 50% or less, we performed DTR. RESULTS: There was no significant difference in the rate of body weight change at 6 or 12 months, or in biochemical markers (hemoglobin, total protein, and albumin) at 12 months. However, 8 patients in the EG group had extreme body weight loss (≥20%) within 12 months. Conversely, in the DTR group, no patients had any extreme body weight loss. CONCLUSION: DTR is useful after laparoscopic proximal gastrectomy, especially in terms of preventing extreme body weight loss.

Postoperative atrial fibrillation does not impact on overall survival after esophagectomy in patients with thoracic esophageal cancer: results from a randomized, double-blind, placebo-controlled trial.

BACKGROUND: Administration of landiolol hydrochloride was found to be associated with reduced incidence of atrial fibrillation (AF) after esophagectomy for esophageal cancer in our previous randomized controlled trial (RCT). In addition, reduced incidence of AF was associated with reduction of other complications. Meanwhile, the effects of postoperative AF and other complications on long-term survival following esophagectomy are not well understood. MATERIALS AND METHODS: Between March 2014 and January 2016, 100 patients with esophageal cancer were registered in an RCT trial and randomly allocated to receive either administration of landiolol or a placebo. We analyzed data from this RCT to better understand the effect of postoperative AF and severe associated complications on overall survival (OS) after esophagectomy for cancer. We also examined whether prophylactic administration of landiolol hydrochloride directly affects prolonged survival in patients with esophageal cancer. RESULTS: The five-year rates of OS in the patients with and without AF were 60%, and 68.6%, respectively, there was no significant difference (P = 0.328). Five-year rates of OS of the patients with and without severe complications were 64.6%, and 67.5%, respectively (P = 0.995). The five-year rates of OS in the placebo and landiolol groups were 65.8% and 68%, respectively (P = 0.809). In multivariate analysis, high stage (stage III/IV) alone was an independent prognostic factor for esophageal cancer patients following esophagectomy. CONCLUSIONS: New-onset AF and the other severe complications were not associated with poorer long-term survival following esophagectomy. In addition, administration of landiolol hydrochloride after esophagectomy did not contribute to prolonging the OS.