Gut microbiota promote liver regeneration through hepatic membrane phospholipid biosynthesis.

BACKGROUND & AIMS: Hepatocyte growth and proliferation depends on membrane phospholipid biosynthesis. Short-chain fatty acids (SCFAs) generated by bacterial fermentation, delivered through the gut-liver axis, significantly contribute to lipid biosynthesis. We therefore hypothesized that dysbiotic insults like antibiotic treatment not only affect gut microbiota, but also impair hepatic lipid synthesis and liver regeneration. METHODS: Stable isotope labeling and 70% partial hepatectomy (PHx) was carried out in C57Bl/6J wild-type mice, in mice treated with broad-spectrum antibiotics, in germ-free mice and mice colonized with minimal microbiota. The microbiome was analyzed by 16S rRNA gene sequencing and microbial culture. Gut content, liver, blood and primary hepatocyte organoids were tested by mass spectrometry-based lipidomics, quantitative reverse-transcription PCR (qRT-PCR), immunoblot and immunohistochemistry for expression of proliferative and lipogenic markers. Matched biopsies from hyperplastic and hypoplastic liver tissue of patients subjected to surgical intervention to induce hyperplasia were analyzed by qRT-PCR for lipogenic enzymes. RESULTS: Three days of antibiotic treatment induced persistent dysbiosis with significantly decreased beta-diversity and richness, but a massive increase of Proteobacteria, accompanied by decreased colonic SCFAs. After PHx, antibiotic-treated mice showed delayed liver regeneration, increased mortality, impaired hepatocyte proliferation and decreased hepatic phospholipid synthesis. Expression of the lipogenic enzyme SCD1 was upregulated after PHx but delayed by antibiotic treatment. Germ-free mice essentially recapitulated the phenotype of antibiotic treatment. Phospholipid biosynthesis, hepatocyte proliferation, liver regeneration and survival were rescued in gnotobiotic mice colonized with a minimal SCFA-producing microbial community. SCFAs induced the growth of murine hepatocyte organoids and hepatic SCD1 expression in mice. Further, SCD1 was required for proliferation of human hepatoma cells and was associated with liver regeneration in human patients. CONCLUSION: Gut microbiota are pivotal for hepatic membrane phospholipid biosynthesis and liver regeneration. IMPACT AND IMPLICATIONS: Gut microbiota affect hepatic lipid metabolism through the gut-liver axis, but the underlying mechanisms are poorly understood. Perturbations of the gut microbiome, e.g. by antibiotics, impair the production of bacterial metabolites, which normally serve as building blocks for membrane lipids in liver cells. As a consequence, liver regeneration and survival after liver surgery is severely impaired. Even though this study is preclinical, its results might allow physicians in the future to improve patient outcomes after liver surgery, by modulation of gut microbiota or their metabolites.

Future directions in preclinical and translational cancer neuroscience research.

Recent advances in cancer neuroscience necessitate the systematic analysis of neural influences in cancer as potential therapeutic targets in oncology. Here, we outline recommendations for future preclinical and translational research in this field.

Patterns and Relevance of Langerhans Islet Invasion in Pancreatic Cancer.

Background: Pancreatic cancer-associated diabetes mellitus (PC-DM) is present in most patients with pancreatic cancer, but its pathogenesis remains poorly understood. Therefore, we aimed to characterize tumor infiltration in Langerhans islets in pancreatic cancer and determine its clinical relevance. METHODS: Langerhans islet invasion was systematically analyzed in 68 patientswith pancreatic ductal adenocarcinoma (PDAC) using histopathological examination and 3D in vitro migration assays were performed to assess chemoattraction of pancreatic cancer cells to isletcells. RESULTS: Langerhans islet invasion was present in all patients. We found four different patterns of islet invasion: (Type I) peri-insular invasion with tumor cells directly touching the boundary, but not penetrating the islet; (Type II) endo-insular invasion with tumor cells inside the round islet; (Type III) distorted islet structure with complete loss of the round islet morphology; and (Type IV)adjacent cancer and islet cells with solitary islet cells encountered adjacent to cancer cells. Pancreatic cancer cells did not exhibit any chemoattraction to islet cells in 3D assays in vitro. Further, there was no clinical correlation of islet invasion using the novel Islet Invasion Severity Score (IISS), which includes all invasion patterns with the occurrence of diabetes mellitus. However, Type IV islet invasion was related to worsened overall survival in our cohort. CONCLUSIONS: We systematically analyzed, for the first time, islet invasion in human pancreatic cancer. Four different main patterns of islet invasion were identified. Diabetes mellitus was not related to islet invasion. However, moreresearch on this prevailing feature of pancreatic cancer is needed to better understand underlying principles.

Clinically Actionable Strategies for Studying Neural Influences in Cancer.

Neuro-glial activation is a recently identified hallmark of growing cancers. Targeting tumor hyperinnervation in preclinical and small clinical trials has yielded promising antitumor effects, highlighting the need of systematic analysis of neural influences in cancer (NIC). Here, we outline the strategies translating these findings from bench to the clinic.

Localisation analysis of nerves in the mouse pancreas reveals the sites of highest nerve density and nociceptive innervation.

BACKGROUND: Neuropathy and neuro-inflammation drive the severe pain and disease progression in human chronic pancreatitis and pancreatic cancer. Mice, especially genetically induced-mouse models, have been increasingly utilized in mechanistic research on pancreatic neuropathy, but the normal "peripheral neurobiology" of the mouse pancreas has not yet been critically compared to human pancreas. METHODS: We introduced a standardized tissue-harvesting technique that preserves the anatomic orientation of the mouse pancreas and allows complete sectioning in an anterior to posterior fashion. We applied immunohistochemistry and quantitative colorimetry of all nerves from the whole organ for studying pancreatic neuro-anatomy. KEY RESULTS: Nerves in the mouse pancreas appeared as "clusters" of nerve trunks in contrast to singly distributed nerve trunks in the human pancreas. Nerve trunks in the mouse pancreas were exclusively found around intrapancreatic blood vessels, and around lymphoid structures. The majority of nerve trunks were located in the pancreatic head (0.15 ± 0.08% of tissue area) and the anterior/front surface of the corpus/body (0.17 ± 0.27%), thus significantly more than in the tail (0.02 ± 0.02%, P = .006). Nerves in the tail included a higher proportion of nociceptive fibers, but the absolute majority, ie, ca. 70%, of all nociceptive fibers, were localized in the head. Mice heterozygous for Bdnf knockout allele (Bdnf+/- ) exhibited enrichment of nitrergic nerve fibers specifically in the head and corpus. CONCLUSIONS & INFERENCES: Neuro-anatomy of the "mesenteric type" mouse pancreas is highly different from the "compact" human pancreas. Studies that aim at reproducing human pancreatic neuro-phenomena in mouse models should pay diligent attention to these anatomic differences.

Neoadjuvant Treatment for Borderline Resectable Pancreatic Ductal Adenocarcinoma.

One of the main reasons for the dismal prognosis of pancreatic ductal adenocarcinoma (PDAC) is its late diagnosis. At the time of presentation, only approximately 15-20% of all patients with PDAC are considered resectable and around 30% are considered borderline resectable. A surgical approach, which is the only curative option, is limited in borderline resectable patients by local involvement of surrounding structures. In borderline resectable pancreatic cancer (BRPC), neoadjuvant treatment regimens have been introduced with the rationale to downstage and downsize the tumor in order to enable resection and eliminate -microscopic distant metastases. However, there are no official guidelines for the preoperative treatment of BRPC. In the majority of cases, patients are administered -Gemcitabine-based or FOLFIRINOX-based chemotherapy regimens with or without radiation. Radiologic restaging after neoadjuvant therapy has to be judged with caution when it comes to predict tumor response and resectability, since inflammation induced by neoadjuvant therapy may mimic solid tumor. Patients who do not show any disease progression during neoadjuvant therapy should be offered surgical exploration, since a high percentage is likely to undergo resection with negative margins (R0) and, thus, achieve improved overall survival although imaging judged it unlikely. Despite the promising new approaches of neoadjuvant treatment regimens during the last 2 decades, surgery remains the first choice if the tumor appears to be primary resectable at the time of diagnosis. At present, there are no international guidelines regarding the preoperative treatment of BRPC. Therefore, in order to standardize and adjust neoadjuvant treatment in the future, new guidelines have to be determined on the basis of upcoming prospective randomized studies.

3D Cancer Migration Assay with Schwann Cells.

In pancreatic cancer, neural invasion is one of the most common paths of cancer dissemination. Classically, cancer cells actively invade nerves and cause local recurrence and pain. Three-dimensional (3D) neural migration assay has become a standard tool for scientists to study neural invasion by confronting the involved cell types. This protocol introduces Schwann cells, i.e., the most prevalent cell type in peripheral nerves, in a novel heterotypic, glia-cancer-neuron, 3D migration assay for assessing their relevance in the early pathogenesis of neural invasion. Particularly, this assay allows the monitoring of the early Schwann cell migratory activity.