Emerging Roles of Spatial Transcriptomics in Liver Research.

Spatial transcriptomics, leveraging sequencing- and imaging-based techniques, has emerged as a groundbreaking technology for mapping gene expression within the complex architectures of tissues. This approach provides an in-depth understanding of cellular and molecular dynamics across various states of healthy and diseased livers. Through the integration of sophisticated bioinformatics strategies, it enables detailed exploration of cellular heterogeneity, transitions in cell states, and intricate cell-cell interactions with remarkable precision. In liver research, spatial transcriptomics has been particularly revelatory, identifying distinct zonated functions of hepatocytes that are crucial for understanding the metabolic and detoxification processes of the liver. Moreover, this technology has unveiled new insights into the pathogenesis of liver diseases, such as the role of lipid-associated macrophages in steatosis and endothelial cell signals in liver regeneration and repair. In the domain of liver cancer, spatial transcriptomics has proven instrumental in delineating intratumor heterogeneity, identifying supportive microenvironmental niches and revealing the complex interplay between tumor cells and the immune system as well as susceptibility to immune checkpoint inhibitors. In conclusion, spatial transcriptomics represents a significant advance in hepatology, promising to enhance our understanding and treatment of liver diseases.

The Spatial Extracellular Proteomic Tumor Microenvironment Distinguishes Molecular Subtypes of Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) mortality rates continue to increase faster than those of other cancer types due to high heterogeneity, which limits diagnosis and treatment. Pathological and molecular subtyping have identified that HCC tumors with poor outcomes are characterized by intratumoral collagenous accumulation. However, the translational and post-translational regulation of tumor collagen, which is critical to the outcome, remains largely unknown. Here, we investigate the spatial extracellular proteome to understand the differences associated with HCC tumors defined by Hoshida transcriptomic subtypes of poor outcome (Subtype 1; S1; n = 12) and better outcome (Subtype 3; S3; n = 24) that show differential stroma-regulated pathways. Collagen-targeted mass spectrometry imaging (MSI) with the same-tissue reference libraries, built from untargeted and targeted LC-MS/MS was used to spatially define the extracellular microenvironment from clinically-characterized, formalin-fixed, paraffin-embedded tissue sections. Collagen α-1(I) chain domains for discoidin-domain receptor and integrin binding showed distinctive spatial distribution within the tumor microenvironment. Hydroxylated proline (HYP)-containing peptides from the triple helical regions of fibrillar collagens distinguished S1 from S3 tumors. Exploratory machine learning on multiple peptides extracted from the tumor regions could distinguish S1 and S3 tumors (with an area under the receiver operating curve of ≥0.98; 95% confidence intervals between 0.976 and 1.00; and accuracies above 94%). An overall finding was that the extracellular microenvironment has a high potential to predict clinically relevant outcomes in HCC.

Robot-assisted transcervical esophagectomy with a bilateral cervical approach for thoracic esophagectomy.

BACKGROUND: Thoracic esophageal cancer resection through the neck approach has recently been reported as mediastinoscopic surgery. We present the first report of a new minimally invasive technique for thoracic esophageal cancer: robot-assisted transcervical esophagectomy with a bilateral cervical approach. METHODS: Ten cases of robot-assisted bilateral transcervical esophagectomy performed at the National Cancer Center Hospital East, Japan, from February 2023 to August 2023 were reviewed. The short-term surgical outcomes were presented, and the feasibility and efficacy of this procedure were discussed. RESULTS: The mean operation time for the cervical procedure was 184.2 ± 23.6 min. The total time for the whole procedure was 472.7 ± 28.4 min, and total intraoperative blood loss was 162.2 ± 40.0 ml. Among the 10 cases, one patient developed recurrent nerve paralysis, one patient developed pulmonary complications, and no patients developed postoperative pneumonia. The median postoperative hospital stay was 22 (range: 12-43) days. No patients developed severe postoperative surgical complications, which were graded as Clavien-Dindo ≥ III. The total number of surgically harvested mediastinal lymph nodes was 37.2 ± 11.2. CONCLUSIONS: Robot-assisted bilateral transcervical esophagectomy, a novel procedure for thoracic esophageal cancer, was safe and feasible. Using this procedure, the incidence of recurrent nerve palsy, which is a problem with transcervical esophagectomy and mediastinoscopic esophagectomy, is expected to decrease.

Total RAMIE with three-field lymph node dissection by a simultaneous two-team approach using a new docking method for esophageal cancer.

BACKGROUND: Thoracic esophageal cancer surgery using robotic approaches for the thoracic and abdominal parts has recently been reported as total robot-assisted minimally invasive esophagectomy (RAMIE). We herein present the first report of a new technique for esophageal cancer: total RAMIE with three-field lymph node dissection (3FLND) by a simultaneous two-team approach using a new docking method. METHODS: We reviewed 20 patients who underwent total RAMIE with 3FLND by a simultaneous two-team approach at the National Cancer Center East Hospital from March 2023 to September 2023. Short-term surgical outcomes and the safety and efficacy of this technique were analyzed. RESULTS: The mean operative time for abdominal surgery with this new docking technique was 135 ± 19.6 min. The total operative time was 488 ± 42.9 min, and the time from the end of abdominal manipulation to the end of surgery was 80.1 ± 15.6 min. The intraoperative blood loss was 116.7 ± 64.4 mL. The incidence of anastomotic leakage, postoperative vocal cord paralysis, and postoperative pneumonia was 10%, 5%, and 10%, respectively. The median postoperative hospital stay was 14 days (range 11-63 days). No in-hospital deaths occurred, and R0 resection was possible in all cases. The average number of lymph nodes dissected was 87.7. CONCLUSION: These results demonstrate that total RAMIE with a simultaneous two-team approach using the new docking method can be safely introduced. The simultaneous cervical and abdominal manipulation with the new docking method allowed total RAMIE without prolonging the operating time, suggesting that it may be a valuable approach for esophageal cancer surgery.

Assessing the outcomes of posterior thoracic para-aortic lymph node dissection after induction chemotherapy in patients with esophageal squamous cell carcinoma.

Posterior thoracic para-aortic lymph node (TPAN) metastasis is a distant metastasis of esophageal cancer. Several case reports have shown that radical esophagectomy and lymphadenectomy for posterior TPAN improve the prognosis of patients with cStage IVB esophageal cancer and solitary posterior TPAN metastasis; however, the true value of this procedure is unclear. The primary objective of this study was to evaluate the short- and long-term outcomes of lymphadenectomy for posterior TPAN after induction chemotherapy in esophageal cancer. This study enrolled 15 patients who underwent radical esophagectomy for cStage IVB esophageal cancer with solitary posterior TPAN metastasis after induction chemotherapy between January 2013 and October 2022 at our hospital. The short- and long-term of radical esophagectomy and lymphadenectomy for posterior TPAN were retrospectively evaluated. All patients who underwent radical esophagectomy and lymphadenectomy for posterior TPAN achieved a pR0 in this study. The median operative time and intraoperative blood loss were 385 minutes and 164 ml, respectively. Four patients (26.7%) had postoperative complications of Clavien-Dindo grade II or more. The median postoperative hospital stay was 15 days. The 5-year overall survival and recurrence-free survival rates were 55.6% (95% confidence interval: 23.1-79.0) and 55.0% (95% confidence interval: 25.3-77.2), respectively. We showed that lymphadenectomy for posterior TPAN metastasis was associated with an improved prognosis of some patients with advanced esophageal cancer. This technique may serve as a viable treatment option for patients who respond well to induction chemotherapy.

Efficiency of conversion surgery for esophageal squamous cell carcinoma with solitary abdominal para-aortic lymph node metastasis.

PURPOSE: Abdominal para-aortic lymph nodes (PANs) are sites of distant metastasis in esophageal squamous cell cancer (ESCC). The prognosis of patients with Stage IVB ESCC and abdominal PAN metastasis is extremely poor. However, chemotherapy for ESCC has recently been developed, and the effectiveness of combined induction therapy and conversion surgery remains unclear. The primary objective of this study was to evaluate the short- and long-term outcomes of conversion surgery for ESCC and solitary abdominal PAN metastases after induction therapy. METHODS: Thirteen patients who underwent conversion esophagectomy for cStage IVB ESCC with solitary abdominal PAN metastasis after induction therapy between January 2017 and October 2022 at our institution were enrolled. The short- and long-term outcomes of conversion surgery were retrospectively evaluated. RESULTS: Three patients (23.1%) had pathological abdominal PAN metastasis, and six patients (46.2%) without pathological abdominal PAN metastasis showed that chemotherapy eliminated the tumors in the abdominal PAN. Three patients (23.1%) had postoperative complications of Clavien-Dindo grade II or higher. The 3-year overall and recurrence-free survival rates were 83.1% and 51.3%, respectively. CONCLUSIONS: Our findings showed that conversion surgery for ESCC and solitary abdominal PAN metastasis led to a good prognosis when induction therapy was successful.

A novel device to assess the oxygen saturation and congestion status of the gastric conduit in thoracic esophagectomy.

BACKGROUND: In thoracic esophagectomy, anastomotic leakage is one of the most important surgical complications. Indocyanine green (ICG) is the most widely used method to assess tissue blood flow; however, this technique has been pointed out to have disadvantages such as difficulty in evaluating the degree of congestion, lack of objectivity in evaluating the degree of staining, and bias easily caused by ICG injection, camera distance, and other factors. Evaluating tissue oxygen saturation (StO2) overcomes these disadvantages and can be performed easily and repeatedly. It is also possible to measure objective values including the degree of congestion. We evaluate novel imaging technology to assess tissue oxygen saturation (StO2) in the gastric conduit during thoracic esophagectomy. METHODS: Fifty patients were enrolled, with seven excluded due to intraoperative findings, leaving 43 for analysis. These patients underwent thoracic esophagectomy for esophageal cancer. The device was used intraoperatively to evaluate tissue oxygen saturation (StO2) and total hemoglobin index (T-HbI), which guided the optimal site for gastric tube anastomosis. The efficacies of StO2 and T-HbI in relation to short-term outcomes were analyzed. RESULTS: StO2, indicating blood supply to the gastric tube, remained stable beyond the right gastroepiploic artery (RGEA) end but significantly decreased distally to the demarcation line (p <  0.05). T-HbI, indicative of congestion, significantly decreased past the RGEA (p <  0.05). Three patients experienced anastomotic leakage. These patients exhibited significantly lower StO2 (p <  0.01) and higher T-HbI (p <  0.01) at both the RGEA end and the demarcation line. Furthermore, the anastomotic site, usually within 3 cm of the RGEA's anorectal side, also showed significantly lower StO2 (p <  0.01) and higher T-HbI (p <  0.01) in patients with anastomotic leakage. CONCLUSIONS: The novel device provides real-time, objective evaluations of blood flow and congestion in the gastric tube. It proves useful for safer reconstruction during thoracic esophagectomy, particularly by identifying optimal anastomosis sites and predicting potential anastomotic leakage.

Effectiveness of 1-year pemafibrate treatment on steatotic liver disease: the influence of alcohol consumption.

BACKGROUND/AIMS: Pemafibrate is a selective peroxisome proliferator-activated receptor α modulator that improves serum alanine aminotransferase (ALT) in dyslipidemia patients. We previously reported that pemafibrate significantly improves liver function, serum triglyceride (TG) levels and liver stiffness in non-alcoholic fatty liver disease patients, however the influence of alcohol consumption was not considered. Therefore, we explored pemafibrate efficacy in patients with steatotic liver disease (SLD) and alcohol-associated liver disease (ALD). METHODS: We retrospectively evaluated pemafibrate efficacy on liver enzymes and lipids in metabolic dysfunction-associated SLD (MASLD) (n = 93), MASLD plus increased alcohol intake (MetALD; n = 23) and ALD (n = 22) patients who had taken pemafibrate for at least 48 weeks. Liver shear wave velocity (SWV, n = 75) was also evaluated. RESULTS: In MASLD group, ALT, aspartate aminotransferase (AST), γ-glutamyl transpeptidase (γ-GTP) and TG values were significantly decreased from baseline to week 24 and week 48 ( P  < 0.0001). ALT and TG values in MetALD group and ALT and AST values in ALD group were also significantly decreased from baseline to week 24 and week 48. Study participant SWV values decreased from baseline to week 48. We observed no significant difference in changes to ALT, AST, γ-GTP and TG (value at week 24 or week 48 minus value at baseline) among the three groups. CONCLUSION: Pemafibrate improves liver function and liver stiffness thus making it a promising therapeutic agent for SLD, even in patients with excess alcohol consumption (MetALD and ALD groups).

Prognosis of patients with residual pathological disease after neoadjuvant docetaxel, cisplatin, and 5-fluorouracil therapy and surgery for esophageal squamous cell carcinoma: a retrospective cohort study.

Background: Docetaxel, cisplatin, and 5-fluorouracil (DCF) combination chemotherapy has been established as one of the standard neoadjuvant therapies for locally advanced esophageal squamous cell carcinoma (ESCC). However, little is known about prognostic factors in patients with residual pathological disease after neoadjuvant DCF followed by surgery for locally advanced ESCC who are candidates for adjuvant nivolumab. Objectives: This study aimed to investigate prognostic factors in patients with residual pathological disease after neoadjuvant DCF chemotherapy followed by surgery for locally advanced ESCC. Design: This was a retrospective cohort study. Methods: This retrospective cohort study included patients who received neoadjuvant DCF followed by surgery for locally advanced ESCC between June 2014 and January 2020 at the National Cancer Center Hospital East. Results: Among a total of 210 patients, 45 patients (21.4%) achieved a pathological complete response. The 3-year disease-free survival (DFS) rate was significantly lower in patients with residual pathological disease than in those with a pathological complete response [53.5% versus 74.5%; hazard ratio (HR): 2.09, 95% confidence interval (CI): 1.16-3.77, p = 0.01]. In patients with residual pathological disease (n = 165), multivariate analysis revealed that pathological node positivity (HR: 3.59, 95% CI: 1.92-6.71, p < 0.01), supraclavicular lymph node metastasis (HR: 2.15, 95% CI: 1.19-3.90, p = 0.01), and lymphovascular invasion (HR: 1.90, 95% CI: 1.14-3.17, p = 0.02) were significantly associated with poor DFS. Conclusion: In this largest-to-date cohort study, patients with residual pathological disease after neoadjuvant DCF followed by surgery for locally advanced ESCC had a poor prognosis. In these patients, pathological node positivity, including supraclavicular lymph node metastasis, and lymphovascular invasion were considered significant prognostic factors.

PKD1 mutant clones within cirrhotic livers inhibit steatohepatitis without promoting cancer.

Somatic mutations in non-malignant tissues are selected for because they confer increased clonal fitness. However, it is uncertain whether these clones can benefit organ health. Here, ultra-deep targeted sequencing of 150 liver samples from 30 chronic liver disease patients revealed recurrent somatic mutations. PKD1 mutations were observed in 30% of patients, whereas they were only detected in 1.3% of hepatocellular carcinomas (HCCs). To interrogate tumor suppressor functionality, we perturbed PKD1 in two HCC cell lines and six in vivo models, in some cases showing that PKD1 loss protected against HCC, but in most cases showing no impact. However, Pkd1 haploinsufficiency accelerated regeneration after partial hepatectomy. We tested Pkd1 in fatty liver disease, showing that Pkd1 loss was protective against steatosis and glucose intolerance. Mechanistically, Pkd1 loss selectively increased mTOR signaling without SREBP-1c activation. In summary, PKD1 mutations exert adaptive functionality on the organ level without increasing transformation risk.