Recent Advances in Pathology: the 2023 Annual Review Issue of The Journal of Pathology.

The 2023 Annual Review Issue of The Journal of Pathology, Recent Advances in Pathology, contains 12 invited reviews on topics of current interest in pathology. This year, our subjects include immuno-oncology and computational pathology approaches for diagnostic and research applications in human disease. Reviews on the tissue microenvironment include the effects of apoptotic cell-derived exosomes, how understanding the tumour microenvironment predicts prognosis, and the growing appreciation of the diverse functions of fibroblast subtypes in health and disease. We also include up-to-date reviews of modern aspects of the molecular basis of malignancies, and our final review covers new knowledge of vascular and lymphatic regeneration in cardiac disease. All of the reviews contained in this issue are written by expert groups of authors selected to discuss the recent progress in their particular fields and all articles are freely available online (https://pathsocjournals.onlinelibrary.wiley.com/journal/10969896). © 2023 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Recent Advances in Pathology: the 2022 Annual Review Issue of The Journal of Pathology.

The 2022 Annual Review Issue of The Journal of Pathology, Recent Advances in Pathology, contains 15 invited reviews on research areas of growing importance in pathology. This year, the articles include those that focus on digital pathology, employing modern imaging techniques and software to enable improved diagnostic and research applications to study human diseases. This subject area includes the ability to identify specific genetic alterations through the morphological changes they induce, as well as integrating digital and computational pathology with 'omics technologies. Other reviews in this issue include an updated evaluation of mutational patterns (mutation signatures) in cancer, the applications of lineage tracing in human tissues, and single cell sequencing technologies to uncover tumour evolution and tumour heterogeneity. The tissue microenvironment is covered in reviews specifically dealing with proteolytic control of epidermal differentiation, cancer-associated fibroblasts, field cancerisation, and host factors that determine tumour immunity. All of the reviews contained in this issue are the work of invited experts selected to discuss the considerable recent progress in their respective fields and are freely available online (https://onlinelibrary.wiley.com/journal/10969896). © 2022 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Recent Advances in Pathology: the 2021 Annual Review Issue of The Journal of Pathology.

The 2021 Annual Review Issue of The Journal of Pathology contains 14 invited reviews on current research areas of particular importance in pathology. The subjects included here reflect the broad range of interests covered by the journal, including both basic and applied research fields but always with the aim of improving our understanding of human disease. This year, our reviews encompass the huge impact of the COVID-19 pandemic, the development and application of biomarkers for immune checkpoint inhibitors, recent advances in multiplexing antigen/nucleic acid detection in situ, the use of genomics to aid drug discovery, organoid methodologies in research, the microbiome in cancer, the role of macrophage-stroma interactions in fibrosis, and TGF-ß as a driver of fibrosis in multiple pathologies. Other reviews revisit the p53 field and its lack of clinical impact to date, dissect the genetics of mitochondrial diseases, summarise the cells of origin and genetics of sarcomagenesis, provide new data on the role of TRIM28 in tumour predisposition, review our current understanding of cancer stem cell niches, and the function and regulation of p63. The reviews are authored by experts in their field from academia and industry, and provide comprehensive updates of the chosen areas, in which there has been considerable recent progress. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Recent Advances in Pathology: the 2020 Annual Review Issue of The Journal of Pathology.

This year's Annual Review Issue of The Journal of Pathology contains 18 invited reviews on current research areas in pathology. The subject areas reflect the broad range of topics covered by the journal and this year encompass the development and application of software in digital histopathology, implementation of biomarkers in pathology practice; genetics and epigenetics, and stromal influences in disease. The reviews are authored by experts in their field and provide comprehensive updates in the chosen areas, in which there has been considerable recent progress in our understanding of disease. © 2020 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Recent Advances in Pathology: the 2019 Annual Review Issue of The Journal of Pathology.

In this Annual Review Issue of The Journal of Pathology, we present 15 invited reviews on topical aspects of pathology, ranging from the impacts of the microbiome in human disease through mechanisms of cell death and autophagy to recent advances in immunity and the uses of genomics for understanding, classifying and treating human cancers. Each of the reviews is authored by experts in their fields and our intention is to provide comprehensive updates in specific areas of pathology in which there has been considerable recent progress. Copyright © 2019 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Recommendations for minimum information for publication of experimental pathology data: MINPEPA guidelines.

Animal models are essential research tools in modern biomedical research, but there are concerns about their lack of reproducibility and the failure of animal data to translate into advances in human medical therapy. A major factor in improving experimental reproducibility is thorough communication of research methodologies. The recently published ARRIVE guidelines outline basic information that should be provided when reporting animal studies. This paper builds on ARRIVE by providing the minimum information needed in reports to allow proper assessment of pathology data gathered from animal tissues. This guidance covers aspects of experimental design, technical procedures, data gathering, analysis, and presentation that are potential sources of variation when creating morphological, immunohistochemical (IHC) or in situ hybridization (ISH) datasets. This reporting framework will maximize the likelihood that pathology data derived from animal experiments can be reproduced by ensuring that sufficient information is available to allow for replication of the methods and facilitate inter-study comparison by identifying potential interpretative confounders.

BCL-3 expression promotes colorectal tumorigenesis through activation of AKT signalling.

OBJECTIVE: Colorectal cancer remains the fourth most common cause of cancer-related mortality worldwide. Here we investigate the role of nuclear factor-κB (NF-κB) co-factor B-cell CLL/lymphoma 3 (BCL-3) in promoting colorectal tumour cell survival. DESIGN: Immunohistochemistry was carried out on 47 tumour samples and normal tissue from resection margins. The role of BCL-3/NF-κB complexes on cell growth was studied in vivo and in vitro using an siRNA approach and exogenous BCL-3 expression in colorectal adenoma and carcinoma cells. The question whether BCL-3 activated the AKT/protein kinase B (PKB) pathway in colorectal tumour cells was addressed by western blotting and confocal microscopy, and the ability of 5-aminosalicylic acid (5-ASA) to suppress BCL-3 expression was also investigated. RESULTS: We report increased BCL-3 expression in human colorectal cancers and demonstrate that BCL-3 expression promotes tumour cell survival in vitro and tumour growth in mouse xenografts in vivo, dependent on interaction with NF-κB p50 or p52 homodimers. We show that BCL-3 promotes cell survival under conditions relevant to the tumour microenvironment, protecting both colorectal adenoma and carcinoma cells from apoptosis via activation of the AKT survival pathway: AKT activation is mediated via both PI3K and mammalian target of rapamycin (mTOR) pathways, leading to phosphorylation of downstream targets GSK-3ß and FoxO1/3a. Treatment with 5-ASA suppressed BCL-3 expression in colorectal cancer cells. CONCLUSIONS: Our study helps to unravel the mechanism by which BCL-3 is linked to poor prognosis in colorectal cancer; we suggest that targeting BCL-3 activity represents an exciting therapeutic opportunity potentially increasing the sensitivity of tumour cells to conventional therapy.

Lineage tracing reveals multipotent stem cells maintain human adenomas and the pattern of clonal expansion in tumor evolution.

The genetic and morphological development of colorectal cancer is a paradigm for tumorigenesis. However, the dynamics of clonal evolution underpinning carcinogenesis remain poorly understood. Here we identify multipotential stem cells within human colorectal adenomas and use methylation patterns of nonexpressed genes to characterize clonal evolution. Numerous individual crypts from six colonic adenomas and a hyperplastic polyp were microdissected and characterized for genetic lesions. Clones deficient in cytochrome c oxidase (CCO(-)) were identified by histochemical staining followed by mtDNA sequencing. Topographical maps of clone locations were constructed using a combination of these data. Multilineage differentiation within clones was demonstrated by immunofluorescence. Methylation patterns of adenomatous crypts were determined by clonal bisulphite sequencing; methylation pattern diversity was compared with a mathematical model to infer to clonal dynamics. Individual adenomatous crypts were clonal for mtDNA mutations and contained both mucin-secreting and neuroendocrine cells, demonstrating that the crypt contained a multipotent stem cell. The intracrypt methylation pattern was consistent with the crypts containing multiple competing stem cells. Adenomas were epigenetically diverse populations, suggesting that they were relatively mitotically old populations. Intratumor clones typically showed less diversity in methylation pattern than the tumor as a whole. Mathematical modeling suggested that recent clonal sweeps encompassing the whole adenoma had not occurred. Adenomatous crypts within human tumors contain actively dividing stem cells. Adenomas appeared to be relatively mitotically old populations, pocketed with occasional newly generated subclones that were the result of recent rapid clonal expansion. Relative stasis and occasional rapid subclone growth may characterize colorectal tumorigenesis.

Epidermal growth factor attenuates tubular necrosis following mercuric chloride damage by regeneration of indigenous, not bone marrow-derived cells.

To assess effects of epidermal growth factor (EGF) and pegylated granulocyte colony-stimulating factor (P-GCSF; pegfilgrastim) administration on the cellular origin of renal tubular epithelium regenerating after acute kidney injury initiated by mercuric chloride (HgCl2 ). Female mice were irradiated and male whole bone marrow (BM) was transplanted into them. Six weeks later recipient mice were assigned to one of eight groups: control, P-GCSF+, EGF+, P-GCSF+EGF+, HgCl2 , HgCl2 +P-GCSF+, HgCl2 +EGF+ and HgCl2 +P-GCSF+EGF+. Following HgCl2 , injection tubular injury scores increased and serum urea nitrogen levels reached uraemia after 3 days, but EGF-treated groups were resistant to this acute kidney injury. A four-in-one analytical technique for identification of cellular origin, tubular phenotype, basement membrane and S-phase status revealed that BM contributed 1% of proximal tubular epithelium in undamaged kidneys and 3% after HgCl2 damage, with no effects of exogenous EGF or P-GCSF. Only 0.5% proximal tubular cells were seen in S-phase in the undamaged group kidneys; this increased to 7-8% after HgCl2 damage and to 15% after addition of EGF. Most of the regenerating tubular epithelium originated from the indigenous pool. BM contributed up to 6.6% of the proximal tubular cells in S-phase after HgCl2 damage, but only to 3.3% after additional EGF. EGF administration attenuated tubular necrosis following HgCl2 damage, and the major cause of this protective effect was division of indigenous cells, whereas BM-derived cells were less responsive. P-GCSF did not influence damage or regeneration.

Fibroblast growth factor receptor (FGFR) gene amplifications are rare events in bladder cancer.

AIMS: Activating point mutations and protein overexpression of fibroblast growth factor receptors (FGFRs), especially FGFR3, are frequent events in bladder cancer. Little is known about gene amplifications, therefore we characterized amplification of FGFR1-3 by fluorescence in-situ hybridization (FISH). METHODS AND RESULTS: Tumours of 153 patients (n = 65 pTa low-grade, n = 15 pTa high-grade, n = 37 pT1, n = 20 pT2, n = 10 pT3, n = 6 pT4) were analysed by FISH for FGFR1-3 copy numbers and screened for FGFR3 mutations and immunohistochemical protein expression. Amplifications of FGFR1 were found in 1.6% (two of 122), FGFR2 in 0.8% (one of 121) and FGFR3 in 3.4% (five of 145). All amplifications were high-level amplifications, not overlapping with polysomy. Amplifications were found in papillary/papillary-invasive tumour parts, and predominantly in tumours with enhanced Ki67 index (>10%), aberrant CK20 expression, and low p53 expression. All FGFR3-amplified samples showed concomitant FGFR3 mutations and FGFR3 protein overexpression. FGFR amplifications were not associated significantly with gender, age, grade or stage in statistical analyses. CONCLUSIONS: FGFR amplifications are rare events in bladder cancer, with FGFR3 amplification being the most prevalent (3.4% of cases). Concomitant FGFR3 mutations and protein overexpression indicate that FGFR3-mediated signalling in these tumours would probably be highly active. This patient subgroup may be particularly suited to FGFR-targeted pharmacotherapy.

Targeted inactivation of fh1 causes proliferative renal cyst development and activation of the hypoxia pathway.

Germline mutations in the fumarate hydratase (FH) tumor suppressor gene predispose to leiomyomatosis, renal cysts, and renal cell cancer (HLRCC). HLRCC tumors overexpress HIF1alpha and hypoxia pathway genes. We conditionally inactivated mouse Fh1 in the kidney. Fh1 mutants developed multiple clonal renal cysts that overexpressed Hif1alpha and Hif2alpha. Hif targets, such as Glut1 and Vegf, were upregulated. We found that Fh1-deficient murine embryonic stem cells and renal carcinomas from HLRCC showed similar overexpression of HIF and hypoxia pathway components to the mouse cysts. Our data have shown in vivo that pseudohypoxic drive, resulting from HIF1alpha (and HIF2alpha) overexpression, is a direct consequence of Fh1 inactivation. Our mouse may be useful for testing therapeutic interventions that target angiogenesis and HIF-prolyl hydroxylation.

The stem cell organisation, and the proliferative and gene expression profile of Barrett's epithelium, replicates pyloric-type gastric glands.

OBJECTIVE: Barrett's oesophagus shows appearances described as 'intestinal metaplasia', in structures called 'crypts' but do not typically display crypt architecture. Here, we investigate their relationship to gastric glands. METHODS: Cell proliferation and migration within Barrett's glands was assessed by Ki67 and iododeoxyuridine (IdU) labelling. Expression of mucin core proteins (MUC), trefoil family factor (TFF) peptides and LGR5 mRNA was determined by immunohistochemistry or by in situ hybridisation, and clonality was elucidated using mitochondrial DNA (mtDNA) mutations combined with mucin histochemistry. RESULTS: Proliferation predominantly occurs in the middle of Barrett's glands, diminishing towards the surface and the base: IdU dynamics demonstrate bidirectional migration, similar to gastric glands. Distribution of MUC5AC, TFF1, MUC6 and TFF2 in Barrett's mirrors pyloric glands and is preserved in Barrett's dysplasia. MUC2-positive goblet cells are localised above the neck in Barrett's glands, and TFF3 is concentrated in the same region. LGR5 mRNA is detected in the middle of Barrett's glands suggesting a stem cell niche in this locale, similar to that in the gastric pylorus, and distinct from gastric intestinal metaplasia. Gastric and intestinal cell lineages within Barrett's glands are clonal, indicating derivation from a single stem cell. CONCLUSIONS: Barrett's shows the proliferative and stem cell architecture, and pattern of gene expression of pyloric gastric glands, maintained by stem cells showing gastric and intestinal differentiation: neutral drift may suggest that intestinal differentiation advances with time, a concept critical for the understanding of the origin and development of Barrett's oesophagus.

In Crohn's disease fibrosis-reduced expression of the miR-29 family enhances collagen expression in intestinal fibroblasts.

Intestinal fibrosis with stricture formation is a complication of CD (Crohn's disease) that may mandate surgical resection. Accurate biomarkers that reflect the relative contribution of fibrosis to an individual stricture are an unmet need in managing patients with CD. The miRNA-29 (miR-29) family has been implicated in cardiac, hepatic and pulmonary fibrosis. In the present study, we investigated the expression of miR-29a, miR-29b and miR-29c in mucosa overlying a stricture in CD patients (SCD) paired with mucosa from non-strictured areas (NSCD). There was significant down-regulation of the miR-29 family in mucosa overlying SCD compared with mucosa overlying NSCD. miR-29b showed the largest fold-decrease and was selected for functional analysis. Overexpression of miR-29b in CD fibroblasts led to a down-regulation of collagen I and III transcripts and collagen III protein, but did not alter MMP (matrix metalloproteinase)-3, MMP-12 and TIMP (tissue inhibitor of metalloproteinase)-1 production. TGF (transforming growth factor)-ß1 up-regulated collagen I and III transcripts and collagen III protein as a consequence of the down-regulation of miR-29b, and TGF-ß1-induced collagen expression was reversed by exogenous overexpression of miR-29b. Furthermore, serum levels of miR-29 were lower in patients with stricturing disease compared with those without. These findings implicate the miR-29 family in the pathogenesis of intestinal fibrosis in CD and provide impetus for the further evaluation of the miR-29 family as biomarkers.

Omnis cellula e cellula revisited: cell biology as the foundation of pathology.

This 2012 Annual Review Issue of The Journal of Pathology argues strongly that cell biology, in its many disciplines, underpins the foundation of our understanding of the mechanisms of disease-the holy grail of pathology. Our increasing knowledge of the human genome will not be enough to attain this goal without parallel developments in our comprehension of the results, at the cellular level, of these genetic changes. In the end, it is cell biology and cell biologists who will deliver this mission.

The C-terminus of Apc does not influence intestinal adenoma development or progression.

Adenomatous polyposis coli (APC ) mutations are found in most colorectal tumours. These mutations are almost always protein-truncating, deleting both central domains that regulate Wnt signalling and C-terminal domains that interact with the cytoskeleton. The importance of Wnt dysregulation for colorectal tumourigenesis is well characterized. It is, however, unclear whether loss of C-terminal functions contributes to tumourigenesis, although this protein region has been implicated in cellular processes--including polarity, migration, mitosis, and chromosomal instability (CIN)­that have been postulated as critical for the development and progression of intestinal tumours. Since almost all APC mutations in human patients disrupt both central and C-terminal regions, we created a mouse model to test the role of the C-terminus of APC in intestinal tumourigenesis. This mouse (Apc(ΔSAMP)) carries an internal deletion within Apc that dysregulates Wnt by removing the beta-catenin-binding and SAMP repeats, but leaves the C-terminus intact. We compared Apc(ΔSAMP) mice with Apc(1322T) animals. The latter allele represented the most commonly found human APC mutation and was identical to Apc(ΔSAMP) except for absence of the entire C-terminus. Apc(ΔSAMP) mice developed numerous intestinal adenomas indistinguishable in number, location, and dysplasia from those seen in Apc(1322T) mice. No carcinomas were found in Apc(ΔSAMP) or Apc(1322T) animals. While similar disruption of the Wnt signalling pathway was observed in tumours from both mice, no evidence of differential C-terminus functions (such as cell migration, CIN, or localization of APC and EB1) was seen. We conclude that the C-terminus of APC does not influence intestinal adenoma development or progression.

CUTL1 is a target of TGF(beta) signaling that enhances cancer cell motility and invasiveness.

CUTL1, also known as CDP, Cut, or Cux-1, is a homeodomain transcriptional regulator known to be involved in development and cell cycle progression. Here we report that CUTL1 activity is associated with increased migration and invasiveness in numerous tumor cell lines, both in vitro and in vivo. Furthermore, we identify CUTL1 as a transcriptional target of transforming growth factor beta and a mediator of its promigratory effects. CUTL1 activates a transcriptional program regulating genes involved in cell motility, invasion, and extracellular matrix composition. CUTL1 expression is significantly increased in high-grade carcinomas and is inversely correlated with survival in breast cancer. This suggests that CUTL1 plays a central role in coordinating a gene expression program associated with cell motility and tumor progression.

Bone marrow-derived cells contribute to cerulein-induced pancreatic fibrosis in the mouse.

Bone marrow (BM) cells may transdifferentiate into circulating fibrocytes and myofibroblasts in organ fibrosis. In this study, we investigated the contribution and functional roles of BM-derived cells in murine cerulein-induced pancreatic fibrosis. C57/BL6 female mice wild-type (WT) or Col 1α1(r/r) male BM transplant, received supraphysiological doses of cerulein to induce pancreatic fibrosis. The CD45(+)Col 1(+) fibrocytes isolated from peripheral blood (PB) and pancreatic tissue were examined by in situ hybridization for Y chromosome detection. The number of BM-derived myofibroblasts, the degree of Sirius red staining and the levels of Col 1α1 mRNA were quantified. The Y chromosome was detected in the nuclei of PB CD45(+)Col 1(+) fibrocytes, confirming that circulating fibrocytes can be derived from BM. Co-expression of α-smooth muscle actin illustrated that fibrocytes can differentiate into myofibroblasts. The number of BM-derived myofibroblasts, degree of collagen deposition and pro-collagen I mRNA expression were higher in the mice that received Col 1α1(r/r) BM, (cells that produce mutated, collagenase-resistant collagen) compared to WT BM, indicating that the genotype of BM cells can alter the degree of pancreatic fibrosis. Our data indicate that CD45(+)Col 1(+) fibrocytes in the PB can be BM-derived, functionally contributing to cerulein-induced pancreatic fibrosis in mice by differentiating into myofibroblasts.

The Journal of Pathology's approach to publication ethics and misconduct.

This Editorial highlights recent changes at The Journal of Pathology intended to improve our ability to detect, and we hope deter, instances of ethical misconduct among submissions made to the Journal, such as cases of guest authorship and plagiarism. We also discuss our experience to date and describe our policies for dealing with such cases. These changes are all encapsulated in our full online Author Guidelines.

Elevation of WNT5A expression in polyp formation in Lkb1+/- mice and Peutz-Jeghers syndrome.

Peutz-Jeghers syndrome (PJS) is a rare, inherited disease caused by germline mutation of the LKB1 gene. Patients with PJS develop characteristic polyps in the digestive tract and carry an elevated risk of cancers in multiple organs, including the intestinal tract. While LKB1 is capable of phosphorylating AMPK and regulates the mTOR pathway, it is also known to be a multitasking protein that can influence other cellular processes, including cell polarity. We hypothesized that there may be other biological pathways directly or indirectly affected by the loss of LKB1 in PJS and aimed to investigate this possibility through transcriptional profiling of polyps harvested from an Lkb1(+/-) mouse model of PJS and from PJS patients. We identified alterations in the mRNA level of a wide range of genes, including some that are involved in Wnt signalling (Wnt5a, Wif1, Dixdc1, Wnt11, Ccnd1, and Ccnd2), although we did not observe nuclear localization of ß-catenin in over 93 human PJS intestinal polyps or in 24 gastric polyps from Lkb1(+/-) mice. Among these genes, WNT5A, a non-canonical and non-transforming Wnt, is consistently up-regulated in both Lkb1(+/-) mice and human PJS polyps at a high level. We performed in situ hybridization to further define the spatial expression pattern of WNT5A and observed a strong signal in the stroma of mouse and human polyps compared to no or very low expression in the mucosa. Our findings indicate that WNT5A plays an important role in PJS polyposis.

Clonal architecture of human prostatic epithelium in benign and malignant conditions.

The location of stem cells in the epithelium of the prostatic acinus remains uncertain, as does the cellular origin of prostatic neoplasia. Here, we apply lineage tracing to visualize the clonal progeny of stem cells in benign and malignant human prostates and understand the clonal architecture of this epithelium. Cells deficient for the mitochondrially-encoded enzyme cytochrome c oxidase (CCO) were identified in 27 frozen prostatectomy specimens using dual colour enzyme histochemistry and individual CCO-normal and -deficient cell areas were laser-capture microdissected. PCR-sequencing of the entire mitochondrial genome (mtDNA) of cells from CCO-deficient areas found to share mtDNA mutations not present in adjacent CCO-normal cells, thus proving a clonal origin. Immunohistochemistry was performed to visualize the three cell lineages normally present in the prostatic epithelium. Entire CCO-deficient acini, and part-deficient acini were found. Deficient patches spanned either basal or luminal cells, but sometimes also both epithelial cell types in normal, hyperplastic or atrophic epithelium, and prostatic intraepithelial neoplasia (PIN). Patches comprising both PIN and invasive cancer were observed. Each cell area within a CCO-deficient patch contained an identical mtDNA mutation, defining the patch as a clonal unit. CCO-deficient patches in benign epithelium contained basal, luminal and endocrine cells, demonstrating multilineage differentiation and therefore the presence of a stem cell. Our results demonstrate that the normal, atrophic, hypertrophic and atypical (PIN) epithelium of human prostate contains stem cell-derived clonal units that actively replenish the epithelium during ageing. These deficient areas usually included the basal compartment indicating the basal layer as the location of the stem cell. Importantly, single clonal units comprised both PIN and invasive cancer, supporting PIN as the pre-invasive lesion for prostate cancer.