熊谷 祐子

BACKGROUND: Low-density granulocytes (LDGs) have been shown to be increased in the peripheral blood of patients with inflammatory and malignant diseases. This study evaluated LDGs in patients who underwent radical surgery for colorectal cancer (CRC) and their impact on survival. METHODS: Patients who underwent radical colectomy between 2017 to 2021 were screened for enrolment in the study. Peripheral blood was obtained in the operating room before and after surgery and cells were recovered from the mononuclear layer after density gradient preparations. The ratio of CD66b(+) LDG to CD45(+) leukocytes was determined with flow cytometry, and the association of the ratios with patient outcomes was examined. The main outcome of interest was recurrence-free survival (RFS). RESULTS: Out of 228 patients treated, 176 were enrolled, including 108 colonic and 68 rectal cancers. Overall, 38 patients were stage I, 30 were stage II, 72 were stage 3, and 36 were stage IV. The number of LDGs was markedly increased immediately after surgery and the proportion of LDGs correlated positively with operating time (r = 0.2806, P < 0.001) and intraoperative blood loss (r = 0.1838, P = 0.014). Purified LDGs produced high amounts of neutrophil extracellular traps after short-term culture and efficiently trapped tumour cells in vitro. The proportion of postoperative LDGs was significantly higher in 13 patients who developed recurrence (median 9 (range 1.63-47.0)) per cent versus median 2.93 ((range 0.035-59.45) per cent, P = 0.013). When cut-off values were set at 4.9 per cent, a higher proportion of LDGs was strongly and independently associated with decreased RFS (P = 0.005). In patients with stage III disease, adjuvant chemotherapy significantly improved RFS of patients with high ratios of LDGs, but not low LDGs. CONCLUSION: LDGs are recruited to circulating blood by surgical stress early in the postoperative interval after colectomy for colonic cancer and their postoperative proportion is correlated with recurrence.

BACKGROUND/AIMS: Recent studies have demonstrated that the populations of several microbes are significantly increased in fecal samples from patients with colorectal cancer (CRC), suggesting their involvement in the development of CRC. The aim of this study was to identify microbes which are increased in distal CRCs and to identify the specific location of microbes increased in mucosal tissue around the tumor. METHODS: Tissue specimens were collected from surgical resections of 28 distal CRCs. Five samples were collected from each specimen (location A: tumor, B: adjacent normal mucosa, C: normal mucosa 1 cm proximal to the tumor, D: normal mucosa 3 cm proximally, and E: normal mucosa 6 cm proximally). The microbiota in the sample were analyzed using 16S rRNA gene amplicon sequencing and the relative abundance (RA) of microbiota compared among the 5 locations. RESULTS: At the genus level, the RA of Fusobacterium and Streptococcus at location A was the highest among the 5 locations, significantly different from that in location E. The dominant species of each genus was Fusobacterium nucleatum and Streptococcus anginosus. The RAs of these species gradually decreased from locations B to E with a statistically significant difference in F. nucleatum. The genus Peptostreptococcus also showed a similar trend, and the RA of Peptostreptococcus stomatis in location A was significantly associated with depth of tumor invasion and tumor size. CONCLUSION: Although the clinical relevance is not clear yet, these results suggest that F. nucleatum, S. anginosus, and P. stomatis can spread to the adjacent normal tissues and may change the surrounding microenvironment to support the progression of CRC.

PURPOSE: The aim of this study was to evaluate both the intestinal mucosa staple line integrity and anastomotic leak pressure after healing in a porcine survival model. METHODS: We used two suture models using two different size staples (incomplete mucosal closure model: group G [staple height 0.75 mm], complete mucosal closure model: group B [staple height 1.5 mm]) in the porcine ileum. Five staple lines were created in each group made in the ileum for each model, and the staple sites harvested on days 0, 2, and 7. The leak pressure at the staple site was measured at each time point. RESULTS: On day 0, the leak pressure for group G (79.5 mmHg) was significantly lower than that for group B (182.3 mmHg) (p < 0.01). On days 2 and 7, there was no significant difference between groups G and B (171 mmHg and 175.5 mmHg on day 2, 175.5 mmHg and 175.5 mmHg on day 7, p > 0.05). The histological findings in both groups showed similar healing at postoperative days 2 and 7. CONCLUSION: The integrity of the mucosal staple lines was associated with the postoperative leak pressure on day 0. However, there was no association with the leak pressure at two days or more postoperatively in a porcine model.

75歳以上の高齢者における腹腔鏡下大腸手術の治療成績を検討したので報告する。当院において2010年1月から2017年6月までの期間に大腸癌に対し腹腔鏡下大腸手術を施行した565例を対象とし、75歳未満の若年者群(474例)と75歳以上の高齢者群(91例)の2群間の短期成績を比較検討した。高齢者群において術前併存症が多くASA scoreは高値であった。また、高齢者群においてD3郭清の施行率が低く(高齢者群41.8%vs若年者群54.0%、p=0.04)、開腹移行率が高値であった(高齢者群4.4%vs若年者群1.1%、p=0.04)。しかし両群間において手術時間、術中出血量、術後在院日数、術後合併症発生率について有意差を認めなかった。以上より75歳以上高齢者においても75歳未満の若年者と同様に腹腔鏡下大腸手術を選択することが可能である。(著者抄録)

BACKGROUND: To explore best practices for acute cholecystitis, it is necessary to construct a system to assess the difficulty of laparoscopic cholecystectomy (LC) based on intraoperative findings. In this study, multiple evaluators assessed videos of LC to assemble a library of typical video clips for 25 intraoperative findings. METHODS: We have previously identified 25 items that contribute to surgical difficulty in LC. For each item, roughly 30-second video clips were submitted from videos of LC performed at member institutions. We then selected one typical video from the collected clips based on simple tabulation of the instances of agreement. Inter-rater agreement was assessed with Fleiss's κ and Gwet's agreement coefficient (AC). RESULTS: Except in the case of two assessment items ("edematous change" and "easy bleeding"), κ or AC significantly exceeded 0.5 and the typical videos were judged to be applicable. For the two remaining items, the evaluation was repeated after clarifying the definitions of positive and negative findings. Eventually, they were recognized as typical. The completed video clip library contains 31 clips and is divided into five categories (http://www.jshbps.jp/modules/project/index.php?content_id=13). CONCLUSIONS: This clip library may be highly useful in clinical settings as a more objective standard for assessing surgical difficulty in LC.

Aim: PD-1/PD-L1 blockade therapy is now widely used for the treatment of advanced malignancies. Although PD-L1 is known to be expressed by various host cells as well as tumor cells, the role of PD-L1 on non-malignant cells and its clinical significance is unknown. We evaluated cell type-specific expression of PD-L1 in colorectal cancer (CRC) specimens using multicolor flow cytometry. Methods: Single cell suspensions were made from 21 surgically resected CRC specimens, and immunostained with various mAbs conjugated with different fluorescent dyes. Tumor cells, stromal cells, and immune cells were identified as CD326(+), CD90(+) and CD45(+) phenotype, respectively. CD11b(+) myeloid cells, CD19(+) B cells and CD4(+) or CD8(+) T cells were also stained in different samples, and their frequencies in the total cell population and the ratio of PD-L1(+) cells to each phenotype were determined. Results: PD-L1 was expressed by all the cell types. The ratio of PD-L1(+) cells to CD326(+) tumor cells was 19.1% ± 14.0%, lower than those for CD90(+) stromal cells (39.6% ± 16.0%) and CD11b(+) myeloid cells (31.9% ± 14.3%). The ratio of PD-L1(+) cells in tumor cells correlated strongly with the ratio in stromal cells, while only weakly with that in myeloid cells. Tumor cells were divided into two populations by CD326 expression levels, and the PD-L1 positive ratios were inversely correlated with the rate of CD326 highly expressing cells as well as mean fluorescein intensity of CD326 in tumor cells, while positively correlated with the frequencies of stromal cells or myeloid cells in CRC. Conclusion: PD-L1 is differentially expressed on various cell types in CRC. PD-L1 on tumor cells may be upregulated together with CD326 downregulation in the process of epithelial mesenchymal transition. Quantification of cell type-specific expression of PD-L1 using multicolor flow cytometry may provide useful information for the immunotherapy of solid tumors.

The peritoneal surface is the most frequent site of metastasis disease in patients with gastric cancer. Even after curative surgery and adjuvant chemotherapy, peritoneal recurrences often develop. Exosomes play pivotal roles in tumor metastasis via the transfer of microRNAs (miRNAs). In the present study, exosomes were isolated from peritoneal lavage fluid or ascites in 85 patients with gastric cancer and the relative expression levels of miR‑29s were examined. The expression of miR‑29a‑3p, miR‑29b‑3p and miR‑29c‑3p in peritoneal exosomes were all downregulated in patients with peritoneal metastases (PM) compared to those without PM. In 30 patients who underwent curative gastrectomy with serosa‑involved (T4) gastric cancer, 6 patients exhibited recurrence in the peritoneum within 12 months. The expression levels of miR‑29s at gastrectomy tended to be lower in these 6 patients than in the other 24 patients with significant differences in miR‑29b‑3p (P=0.003). When the patients were divided into two groups based on median levels of miR‑29s, peritoneal recurrence developed more frequently in patients with low expression of miR‑29b‑3p, and lower expression of miR‑29s were related with worse overall survival. miR‑29s are thought to play a suppressive role in the growth of disseminated peritoneal tumor cells. Reduced expression of miR‑29b in peritoneal exosomes is a strong risk factor of developing postoperative peritoneal recurrence.

BACKGROUND: Low-density neutrophils (LDN) have been shown to be increased in peripheral blood in patients with various diseases and closely related to immune-mediated pathology. However, the frequency and function of LDN in circulating blood of the patients following abdominal surgery have not been well understood. METHODS: LDN were determined by CD66b(+) cells, which were copurified with mononuclear cells by density gradient preparations of peripheral blood of surgical patients. The effects of the purified LDN on T cell proliferation and tumor cell lysis were examined in vitro. Neutrophil extracellular traps (NETs) production was examined by extracellular nuclear staining. RESULTS: The number of LDN with an immature phenotype is markedly increased in peripheral blood samples in patients after abdominal surgery. The frequency of LDN correlated positively with operative time and intraoperative blood loss. The purified LDN significantly suppressed the proliferation of autologous T cells stimulated with anti-CD3 mAb coated on plate and partially inhibited the cytotoxicity of lymphocytes activated with recombinant interleukin-2 against a human gastric cancer cell, OCUM-1. The LDN also produced NETs after short-term culture in vitro, which efficiently trap many OCUM-1. These results suggest that surgical stress recruits immunosuppressive LDN in the circulation in the early postoperative period. CONCLUSIONS: The LDN may support the lodging of circulating tumor cells via NETs formation and inhibit T cell-mediated antitumor response in target organs, which may promote postoperative cancer metastases. Functional blockade of LDN might be an effective strategy to reduce tumor recurrence after abdominal surgery.

Aim: Peritoneal metastases (PM) frequently occur in patients with gastric cancer and result in a poor prognosis. Exosomes play pivotal roles in tumor metastasis through the transfer of microRNAs (miRNAs). We examined the exosomal miRNA profile in peritoneal fluids to identify novel biomarkers to reflect tumor burden in the peritoneum. Methods: Exosomes were isolated from peritoneal fluids of patients of gastric cancer with macroscopic (P1) or microscopic (P0CY1) peritoneal metastasis (PM) and comprehensive miRNA expression analysis was carried out. Expressions of candidate miRNAs were then validated in all 58 samples using TaqMan Advanced miRNA Assays. Results: In initial screening, we carried out comprehensive analysis of exosomal miRNA using peritoneal fluids from 11 and 14 patients with or without PM, respectively, and identified 11 dysregulated miRNAs in PM (+) samples. Validation analysis showed that four miRNAs (miR-21-5p, miR-92a-3p, miR-223-3p, and miR-342-3p) were significantly upregulated in 12 PM (+) samples, and their expression levels showed positive correlation with peritoneal cancer index. In contrast, miR-29 family were all downregulated in patients with PM (+) samples. Moreover, in 24 patients with pT4 tumor, miR-29 at gastrectomy tended to be lower in six patients with peritoneal recurrence with significant differences in miR-29b-3p (P = .012). Conclusion: Expression pattern of miRNAs in peritoneal exosomes well reflects the tumor burden in the peritoneal cavity and could be a useful biomarker in the treatment of PM.

50歳代女。夕食に鯛を摂取し、夜間に腹痛が出現したため当院を受診した。造影CTでは明らかな異常所見がなく、いったん帰宅したが、その後も症状が持続し、再診時のCTで魚骨による食道異物が判明したため、緊急入院となった。内視鏡検査で食道壁に刺入した魚骨を確認し、内視鏡摘出を試みるも困難であったが、1ヵ月後の内視鏡施行時に魚骨の摘出が可能であった。摘出後は出血や穿孔もなく、良好に経過している。