Histological diagnosis of polyploidy discriminates an aggressive subset of hepatocellular carcinomas with poor prognosis.

BACKGROUND: Although genome duplication, or polyploidization, is believed to drive cancer evolution and affect tumor features, its significance in hepatocellular carcinoma (HCC) is unclear. We aimed to determine the characteristics of polyploid HCCs by evaluating chromosome duplication and to discover surrogate markers to discriminate polyploid HCCs. METHODS: The ploidy in human HCC was assessed by fluorescence in situ hybridization for multiple chromosomes. Clinicopathological and expression features were compared between polyploid and near-diploid HCCs. Markers indicating polyploid HCC were explored by transcriptome analysis of cultured HCC cells. RESULTS: Polyploidy was detected in 36% (20/56) of HCCs and discriminated an aggressive subset of HCC that typically showed high serum alpha-fetoprotein, poor differentiation, and poor prognosis compared to near-diploid HCCs. Molecular subtyping revealed that polyploid HCCs highly expressed alpha-fetoprotein but did not necessarily show progenitor features. Histological examination revealed abundant polyploid giant cancer cells (PGCCs) with a distinct appearance and frequent macrotrabecular-massive architecture in polyploid HCCs. Notably, the abundance of PGCCs and overexpression of ubiquitin-conjugating enzymes 2C indicated polyploidy in HCC and efficiently predicted poor prognosis in combination. CONCLUSIONS: Histological diagnosis of polyploidy using surrogate markers discriminates an aggressive subset of HCC, apart from known HCC subgroups, and predict poor prognosis in HCC.

The 6th international cell senescence association conference.

The 6th conference of the international cell senescence association (ICSA) on the theme of "A New Era of Senescence Research: The Challenge of Controlling Aging and Cancer" was held on December 12-15, 2021 in Osaka, Japan as a Hybrid Meeting. The conference brought together basic and translational scientists to discuss recent developments in the field of cellular senescence research. In recent years, the study of cellular senescence has become a very hot field of research. It is clear that the ICSA, founded in 2015, has played an important role in this process. The 6th ICSA conference has provided another opportunity for exchanges and new connections between basic and translational scientists. The scientific program consisted of keynote lectures, invited talks, short talks selected from abstracts, a poster session, and luncheon seminars sponsored by the Japanese Society of Anti-Aging Medicine. In the Meet the Editor session, Dr Christoph Schmitt, Editor-in-Chief of Nature Metabolism, gave a short presentation about the journal and answered questions from the audience. Being a hybrid meeting, there was only so much that could be done, but we hope that the meeting was fruitful.

Implications of Polyploidy and Ploidy Alterations in Hepatocytes in Liver Injuries and Cancers.

Polyploidy, a condition in which more than two sets of chromosomes are present in a cell, is a characteristic feature of hepatocytes. A significant number of hepatocytes physiologically undergo polyploidization at a young age. Polyploidization of hepatocytes is enhanced with age and in a diseased liver. It is worth noting that polyploid hepatocytes can proliferate, in marked contrast to other types of polyploid cells, such as megakaryocytes and cardiac myocytes. Polyploid hepatocytes divide to maintain normal liver homeostasis and play a role in the regeneration of the damaged liver. Furthermore, polyploid hepatocytes have been shown to dynamically reduce ploidy during liver regeneration. Although it is still unclear why hepatocytes undergo polyploidization, accumulating evidence has revealed that alterations in the ploidy in hepatocytes are involved in the pathophysiology of liver cirrhosis and carcinogenesis. This review discusses the significance of hepatocyte ploidy in physiological liver function, liver injury, and liver cancer.

Genetic features of multicentric/multifocal intramucosal gastric carcinoma.

Chronic gastritis caused by Helicobacter pylori (H. pylori) infection could lead to the development of gastric cancer. The finding that multiple gastric cancers can develop synchronously and/or metachronously suggests the development of field cancerization in chronically inflamed, H. pylori-infected gastric mucosa. The genetic basis of multiple tumorigenesis in the inflamed stomach, however, is not well understood. In this study, we analyzed the microsatellite instability (MSI) status and copy number aberrations (CNAs) of 41 multiple intramucosal early gastric cancers that synchronously or metachronously developed in 19 patients with H. pylori infection. Among the 41 intramucosal gastric carcinomas, 9 (22%) exhibited MSI, and the remaining 32 (78%) exhibited the microsatellite stable (MSS) phenotype. Metachronous multiple intramucosal gastric carcinoma exhibit inter-tumor heterogeneity by individually acquiring genetic aberrations. All synchronous multiple intramucosal gastric carcinoma pairs shared a common MSI/MSS profile, and CNA analysis revealed that synchronous multiple intramucosal gastric carcinoma pairs with the MSS phenotype shared common aberrations of representative tumor-suppressor genes, including focal deletion of APC, TP53, CDKN2A, and CDKN2B. Multiregional CNA analysis revealed that heterogeneous gene amplifications/deletions, including PDL1 amplification, evolved under the presence of shared "trunk" genetic alterations in a subpopulation of individual intramucosal gastric carcinomas. These data suggest that multiple gastric carcinomas develop in a multicentric/multifocal manner exhibiting features of inter- and intra-tumor heterogeneity in H. pylori-infected gastric mucosa, whereas synchronous multiple intramucosal gastric carcinomas could share partially common genetic alterations, possibly via common oncogenic pathways.

Stress enhances gait disturbance induced by lumbar disc degeneration in rat.

PURPOSE: Although psychological factors are assumed to be the primary cause of stress-related back pain, there have been no studies of the relationships between stress and low back pain in an animal model. The purpose of this study was to examine the influence of specific alternation of rhythm in temperature (SART) stress on gait abnormality using the CatWalk method in a rat model of low back pain caused by lumbar facetectomy. METHODS: Sixty rats were divided into three groups: the control, sham and experimental groups. Each group was then divided into non-SART stress and SART stress subgroups. We evaluated the behavioral changes 7 weeks postoperatively using the von Frey test and the CatWalk method. RESULTS: Threshold values for the hind paw in the SART stress subgroups were significantly lower than those in the non-SART stress subgroups. In the experimental group, significant changes by CatWalk in step cycle, stand time and average speed were observed under non-SART stress conditions, but SART stress resulted in additional significant changes in not only these parameters, but in other parameters including the duty cycle and swing time, compared with those in the control and sham groups. CONCLUSIONS: The demonstration by CatWalk analysis may indicate that SART stress enhanced gait disturbance. In this animal model, we demonstrated for the first time that stress may be a factor involved in worsening of low back pain.

Hepatic inflammation facilitates transcription-associated mutagenesis via AID activity and enhances liver tumorigenesis.

Chronic inflammation triggers the aberrant expression of a DNA mutator enzyme, activation-induced cytidine deaminase (AID), and contributes to tumorigenesis through the accumulation of genetic aberrations. To gain further insight into the inflammation-mediated genotoxic events required for carcinogenesis, we examined the role of chronic inflammation in the emergence of genetic aberrations in the liver with constitutive AID expression. Treatment with thioacetamide (TAA) at low-dose concentrations caused minimal hepatic inflammation in both wild-type (WT) and AID transgenic (Tg) mice. None of the WT mice with low-dose TAA administration or AID Tg mice without hepatic inflammation developed cancers in their liver tissues over the 6 month study period. In contrast, all the AID Tg mice with TAA treatment developed multiple macroscopic hepatocellular carcinomas during the same observation period. Whole exome sequencing and additional deep-sequencing analyses revealed the enhanced accumulation of somatic mutations in various genes, including dual specificity phosphatase 6 (Dusp6), early growth response 1 (Egr1) and inhibitor of DNA binding 2 (Id2), which are putative tumor suppressors, in AID-expressing liver with TAA-mediated hepatic inflammation. Microarray and quantitative reverse transcription-polymerase chain reaction analyses showed the transcriptional upregulation of various genes including Dusp6, Egr1 and Id2 under hepatic inflammatory conditions. Together, these findings suggest that inflammation-mediated transcriptional upregulation of target genes, including putative tumor suppressor genes, enhances the opportunity for inflamed cells to acquire somatic mutations and contributes to the acceleration of tumorigenesis in the inflamed liver tissues.

Outcomes and Prognostic Factors in Patients with Spontaneously Ruptured Hepatocellular Carcinoma.

BACKGROUND/AIMS: Spontaneous rupture is a life-threatening complication of hepatocellular carcinoma (HCC). Although hemostasis can be achieved by transarterial embolization (TAE), the prognosis remains poor. The aims of this study were to evaluate the effectiveness of emergent TAE for ruptured HCC and to clarify the prognostic factors. METHODOLOGY: Thirty-six patients with spontaneously ruptured HCC were retrospectively analyzed. Prognostic factors of short-term (57 days) and long-term (>7 days) survival after HCC rupture were investigated by univariate and multivariate analyses. RESULTS: Emergent TAE was performed in 22 patients and conservative treatment was applied in 14. The hemostasis rate of TAE was 86.4%, and median survival time in patients with TAE was significantly longer than that in patients with conservative treatment (142 days vs. 5 days, p = 0.0006). In multivariate analysis, high serum creatinine (p = 0.036) was a significant independent predictor of poor 7-day survival, and low serum albumin (p = 0.050) and absence of emergent TAE (p = 0.061) tended to be associated with poor 7-day survival. HCC treatment within the past 12 months (p = 0.048) and, high serum total bilirubin (p = 0.016) were predictors of poor long-term survival. Conclusions: We identified some survival predictors after HCC rupture. Emergent TAE appears to be effective for improving short-term oroLnosis after HCC ruoture.

Impact of intratumoral microbiome on tumor immunity and prognosis in human pancreatic ductal adenocarcinoma.

BACKGROUND: Recent evidence suggests that the presence of microbiome within human pancreatic ductal adenocarcinoma (PDAC) tissue potentially influences cancer progression and prognosis. However, the significance of tumor-resident microbiome remains unclear. We aimed to elucidate the impact of intratumoral bacteria on the pathophysiology and prognosis of human PDAC. METHODS: The presence of intratumoral bacteria was assessed in 162 surgically resected PDACs using quantitative polymerase chain reaction (qPCR) and in situ hybridization (ISH) targeting 16S rRNA. The intratumoral microbiome was explored by 16S metagenome sequencing using DNA extracted from formalin-fixed paraffin-embedded tissues. The profile of intratumoral bacteria was compared with clinical information, pathological findings including tumor-infiltrating T cells, tumor-associated macrophage, fibrosis, and alterations in four main driver genes (KRAS, TP53, CDKN2A/p16, SMAD4) in tumor genomes. RESULTS: The presence of intratumoral bacteria was confirmed in 52 tumors (32%) using both qPCR and ISH. The 16S metagenome sequencing revealed characteristic bacterial profiles within these tumors, including phyla such as Proteobacteria and Firmicutes. Comparison of bacterial profiles between cases with good and poor prognosis revealed a significant positive correlation between a shorter survival time and the presence of anaerobic bacteria such as Bacteroides, Lactobacillus, and Peptoniphilus. The abundance of these bacteria was correlated with a decrease in the number of tumor-infiltrating T cells positive for CD4, CD8, and CD45RO. CONCLUSIONS: Intratumoral infection of anaerobic bacteria such as Bacteroides, Lactobacillus, and Peptoniphilus is correlated with the suppressed anti-PDAC immunity and poor prognosis.

Bacterial induction of B cell senescence promotes age-related changes in the gut microbiota.

The elucidation of the mechanisms of ageing and the identification of methods to control it have long been anticipated. Recently, two factors associated with ageing-the accumulation of senescent cells and the change in the composition of gut microbiota-have been shown to play key roles in ageing. However, little is known about how these phenomena occur and are related during ageing. Here we show that the persistent presence of commensal bacteria gradually induces cellular senescence in gut germinal centre B cells. Importantly, this reduces both the production and diversity of immunoglobulin A (IgA) antibodies that target gut bacteria, thereby changing the composition of gut microbiota in aged mice. These results have revealed the existence of IgA-mediated crosstalk between the gut microbiota and cellular senescence and thus extend our understanding of the mechanism of gut microbiota changes with age, opening up possibilities for their control.

Cellular senescence in white matter microglia is induced during ageing in mice and exacerbates the neuroinflammatory phenotype.

Cellular senescence, a state of irreversible cell-cycle arrest caused by a variety of cellular stresses, is critically involved in age-related tissue dysfunction in various organs. However, the features of cells in the central nervous system that undergo senescence and their role in neural impairment are not well understood as yet. Here, through comprehensive investigations utilising single-cell transcriptome analysis and various mouse models, we show that microglia, particularly in the white matter, undergo cellular senescence in the brain and spinal cord during ageing and in disease models involving demyelination. Microglial senescence is predominantly detected in disease-associated microglia, which appear in ageing and neurodegenerative diseases. We also find that commensal bacteria promote the accumulation of senescent microglia and disease-associated microglia during ageing. Furthermore, knockout of p16INK4a, a key senescence inducer, ameliorates the neuroinflammatory phenotype in damaged spinal cords in mice. These results advance our understanding of the role of cellular senescence in the central nervous system and open up possibilities for the treatment of age-related neural disorders.

Excessive activity of apolipoprotein B mRNA editing enzyme catalytic polypeptide 2 (APOBEC2) contributes to liver and lung tumorigenesis.

Apolipoprotein B mRNA editing enzyme catalytic polypeptide 2 (APOBEC2) was originally identified as a member of the cytidine deaminase family with putative nucleotide editing activity. To clarify the physiologic and pathologic roles, and the target nucleotide of APOBEC2, we established an APOBEC2 transgenic mouse model and investigated whether APOBEC2 expression causes nucleotide alterations in host DNA or RNA sequences. Sequence analyses revealed that constitutive expression of APOBEC2 in the liver resulted in significantly high frequencies of nucleotide alterations in the transcripts of eukaryotic translation initiation factor 4 gamma 2 (Eif4g2) and phosphatase and tensin homolog (PTEN) genes. Hepatocellular carcinoma developed in 2 of 20 APOBEC2 transgenic mice at 72 weeks of age. In addition, constitutive APOBEC2 expression caused lung tumors in 7 of 20 transgenic mice analyzed. Together with the fact that the proinflammatory cytokine tumor necrosis factor-α induces ectopic expression of APOBEC2 in hepatocytes, our findings indicate that aberrant APOBEC2 expression causes nucleotide alterations in the transcripts of the specific target gene and could be involved in the development of human hepatocellular carcinoma through hepatic inflammation.

Development and evaluation of a colorectal cancer screening method using machine learning-based gut microbiota analysis.

Accumulating evidence indicates that alterations of gut microbiota are associated with colorectal cancer (CRC). Therefore, the use of gut microbiota for the diagnosis of CRC has received attention. Recently, several studies have been conducted to detect the differences in the gut microbiota between healthy individuals and CRC patients using machine learning-based gut bacterial DNA meta-sequencing analysis, and to use this information for the development of CRC diagnostic model. However, to date, most studies had small sample sizes and/or only cross-validated using the training dataset that was used to create the diagnostic model, rather than validated using an independent test dataset. Since machine learning-based diagnostic models cause overfitting if the sample size is small and/or an independent test dataset is not used for validation, the reliability of these diagnostic models needs to be interpreted with caution. To circumvent these problems, here we have established a new machine learning-based CRC diagnostic model using the gut microbiota as an indicator. Validation using independent test datasets showed that the true positive rate of our CRC diagnostic model increased substantially as CRC progressed from Stage I to more than 60% for CRC patients more advanced than Stage II when the false positive rate was set around 8%. Moreover, there was no statistically significant difference in the true positive rate between samples collected in different cities or in any part of the colorectum. These results reveal the possibility of the practical application of gut microbiota-based CRC screening tests.

SARS-CoV-2 infection triggers paracrine senescence and leads to a sustained senescence-associated inflammatory response.

Reports of post-acute COVID-19 syndrome, in which the inflammatory response persists even after SARS-CoV-2 has disappeared, are increasing1, but the underlying mechanisms of post-acute COVID-19 syndrome remain unknown. Here, we show that SARS-CoV-2-infected cells trigger senescence-like cell-cycle arrest2,3 in neighboring uninfected cells in a paracrine manner via virus-induced cytokine production. In cultured human cells or bronchial organoids, these SASR-CoV-2 infection-induced senescent cells express high levels of a series of inflammatory factors known as senescence-associated secretory phenotypes (SASPs)4 in a sustained manner, even after SARS-CoV-2 is no longer detectable. We also show that the expression of the senescence marker CDKN2A (refs. 5,6) and various SASP factor4 genes is increased in the pulmonary cells of patients with severe post-acute COVID-19 syndrome. Furthermore, we find that mice exposed to a mouse-adapted strain of SARS-CoV-2 exhibit prolonged signs of cellular senescence and SASP in the lung at 14 days after infection when the virus was undetectable, which could be substantially reduced by the administration of senolytic drugs7. The sustained infection-induced paracrine senescence described here may be involved in the long-term inflammation caused by SARS-CoV-2 infection.

[Signet ring cell carcinoma of the bile duct: a case report].

A 72-year-old man was admitted with obstructive jaundice. Computed tomography revealed a 4cm tumor with multiple cystic components obstructing the common bile duct. Endoscopic ultrasonography, endoscopic retrograde cholangiopancreatography and intraductal ultrasonography demonstrated the tumor, which derived from the lower bile duct, grew into the bile duct lumen. Peroral cholangioscopy revealed distended tumor vessels on the surface of the tumor. Signet ring cell carcinoma of the bile duct was diagnosed by biopsy. The patient died 3 months after the first hospital admission despite chemotherapy.

Proliferative polyploid cells give rise to tumors via ploidy reduction.

Polyploidy is a hallmark of cancer, and closely related to chromosomal instability involved in cancer progression. Importantly, polyploid cells also exist in some normal tissues. Polyploid hepatocytes proliferate and dynamically reduce their ploidy during liver regeneration. This raises the question whether proliferating polyploids are prone to cancer via chromosome missegregation during mitosis and/or ploidy reduction. Conversely polyploids could be resistant to tumor development due to their redundant genomes. Therefore, the tumor-initiation risk of physiologic polyploidy and ploidy reduction is still unclear. Using in vivo lineage tracing we here show that polyploid hepatocytes readily form liver tumors via frequent ploidy reduction. Polyploid hepatocytes give rise to regenerative nodules with chromosome aberrations, which are enhanced by ploidy reduction. Although polyploidy should theoretically prevent tumor suppressor loss, the high frequency of ploidy reduction negates this protection. Importantly, polyploid hepatocytes that undergo multiple rounds of cell division become predominantly mononucleated and are resistant to ploidy reduction. Our results suggest that ploidy reduction is an early step in the initiation of carcinogenesis from polyploid hepatocytes.

Gut bacteria identified in colorectal cancer patients promote tumourigenesis via butyrate secretion.

Emerging evidence is revealing that alterations in gut microbiota are associated with colorectal cancer (CRC). However, very little is currently known about whether and how gut microbiota alterations are causally associated with CRC development. Here we show that 12 faecal bacterial taxa are enriched in CRC patients in two independent cohort studies. Among them, 2 Porphyromonas species are capable of inducing cellular senescence, an oncogenic stress response, through the secretion of the bacterial metabolite, butyrate. Notably, the invasion of these bacteria is observed in the CRC tissues, coinciding with the elevation of butyrate levels and signs of senescence-associated inflammatory phenotypes. Moreover, although the administration of these bacteria into ApcΔ14/+ mice accelerate the onset of colorectal tumours, this is not the case when bacterial butyrate-synthesis genes are disrupted. These results suggest a causal relationship between Porphyromonas species overgrowth and colorectal tumourigenesis which may be due to butyrate-induced senescence.

In Vivo Lineage Tracing of Polyploid Hepatocytes Reveals Extensive Proliferation during Liver Regeneration.

The identity of cellular populations that drive liver regeneration after injury is the subject of intense study, and the contributions of polyploid hepatocytes to organ regeneration and homeostasis have not been systematically assessed. Here, we developed a multicolor reporter allele system to genetically label and trace polyploid cells in situ. Multicolored polyploid hepatocytes undergo ploidy reduction and subsequent re-polyploidization after transplantation, providing direct evidence of the hepatocyte ploidy conveyor model. Marker segregation revealed that ploidy reduction rarely involves chromosome missegregation in vivo. We also traced polyploid hepatocytes in several different liver injury models and found robust proliferation in all settings. Importantly, ploidy reduction was seen in all injury models studied. We therefore conclude that polyploid hepatocytes have extensive regenerative capacity in situ and routinely undergo reductive mitoses during regenerative responses.