Using MsfNet to Predict the ISUP Grade of Renal Clear Cell Carcinoma in Digital Pathology Images

Clear cell renal cell carcinoma(ccRCC)represents the most frequent form of renal cell carcinoma(RCC),and accurate International Society of Urological Pathology(ISUP)grading is crucial for prognosis and treatment selection.This study presents a new deep network called Multi-scale Fusion Network(MsfNet),which aims to enhance the automatic ISUP grade of ccRCC with digital histopathology pathology images.The MsfNet overcomes the limitations of traditional ResNet50 by multi-scale information fusion and dynamic allocation of channel quantity.The model was trained and tested using 90 Hematoxylin and Eosin(H&E)stained whole slide images(WSIs),which were all cropped into 320×320-pixel patches at 40×magnification.MsfNet achieved a micro-averaged area under the curve(AUC)of 0.9807,a macro-averaged AUC of 0.9778 on the test dataset.The Gradient-weighted Class Activation Mapping(Grad-CAM)visually demonstrated MsfNet’s ability to distinguish and highlight abnormal areas more effectively than ResNet50.The t-Distributed Stochastic Neighbor Embedding(t-SNE)plot indicates our model can efficiently extract critical features from images,reducing the impact of noise and redundant information.The results suggest that MsfNet offers an accurate ISUP grade of ccRCC in digital images,emphasizing the potential of AI-assisted histopathological systems in clinical practice.

Microvascular structural changes in esophageal squamous cell carcinoma pathology according to intrapapillary capillary loop types under magnifying endoscopy

BACKGROUND The intrapapillary capillary loop(IPCL)characteristics,visualized using magnifying endoscopy,are commonly assessed for preoperative evaluation of the infiltration depth of esophageal squamous cell carcinoma(ESCC).Japan Esophageal Society(JES)classification is the most widely used classification.Microvascular structural changes are evaluated by magnifying endoscopy for the presence or absence of each morphological factor:tortuosity,dilatation,irregular caliber,and different shapes.However,the pathological characteristics of IPCLs have not been thoroughly investigated,especially the microvascular structures corresponding to the deepest parts of the lesions'infiltration.AIM To investigate differences in pathological microvascular structures of ESCC,which correspond to the deepest parts of the lesions'infiltration.METHODS Patients with ESCC and precancerous lesions diagnosed at Peking University Third Hospital were enrolled between January 2019 and April 2023.Patients first underwent magnified endoscopic examination,followed by endoscopic submucosal dissection or surgical treatment.Pathological images were scanned using a threedimensional slice scanner,and the pathological structural differences in different types,according to the JES classification,were analyzed using nonparametric tests and t-tests.RESULTS The 35 lesions were divided into four groups according to the JES classification:A,B1,B2,and B3.Statistical analyses revealed significant differences(aP<0.05)in the short and long calibers,area,location,and density between types A and B.Notably,there were no significant differences in these parameters between types B1 and B2 and between types B2 and B3(P>0.05).However,significant differences in the short calibers,long calibers,and area of IPCL were observed between types B1 and B3(aP<0.05);no significant differences were found in the density or location(P>0.05).CONCLUSION Pathological structures of IPCLs in the deepest infiltrating regions differ among various IPCL types classified by the JES classification under magnifying endoscopy,especially between the types A and B.

Ultrastructural Pathology of Nerve Cells in Mouse Brain Infected with Epidemic B Encephalitis Virus

An animal model of epidemic(Japanese)B encephalitis was estabilisged by injecting theJin Wei Yah 1 stain of B encephalitis virus into the peritoneal cavity of mice.The ultrastructuralchanges of the nerve cells in their brains were studied,special attention being paid to some types ofnerve cell in the cerebellar cortex.The infectet Purkinje cells and especially the granular cells showedspecial and inter,sting pathological features.These were compared with the changes found in the in-fected nerve cells in the cerebral cortex,diencephalon and mesencephalon.A radiating structure consisting of a microveside-microtubule aggregation body at the centerand endoplastic reticulum or virus replication multivesicular structures around it was often found in thein fected nerve cells.Its morphological features were described in detail,and its significance and the se-quenoe of events of its development discussed.In the late stage of infection,virus particles were found in the nuclei of part of the necroticcells.It is considered that they entered the nuclei from the cytoplasm during or after the death of theinfected cells.The observation smade in this study have comfimed in the granular cell of the cerebellum theidea of Chert et al.that the encephalitis B virus particle can he formed in the perinudear cistem ofthe infected nerve cell,and have brought forth further information in this respect.The way of releaseof the virus particles from the infected nerve cells observed in this study is fundamentally consistentwith that observed by Chen et al.but most of the virus particles left the nerve cell via the cell pro-

Quantitative ultrastucture analysis of neuroendocrine cells of gastric mucosa on normal and pathological conditions

AIMS To study the quantitative ultrastucture of neu- roendocrine cells of gastric mucosa on normal anc pathological conditions including the duodenal ulcer (DU) and Zollinger-Ellison syndrome (ZES). METHODS The neuroendocrine cells of the gastric mucosa of eight normal subjects,six patients with DU and five patients with ZES were quantitatively investi- gated with electro microscope and ultrastructure image analyzer. RESULTS The volume density of neuroendocrine cells in DU was 1.3% and 0.8% (vs 1.6% and 0.9%,P>0.05) in gastric antrum and corpus respectively. In antrum,G cells was of 65% (P< 0.05),D cells decreased in cell density (3% vs 9.5%) and in number of cell per unit area (P<0.01). In corpus,the cell density of ECL cells increased (49% vs 30%,P<0.05);D cells and EC cells decreasec (2% P<0.01 and 4% P<0.05,respectively),and the number of D cell per unit area markedly decreased. In ZES,D cells in corpus decreased in cell density (4% vs 22%,P<0.01) and P cells also decreased (11% vs 24%,P<0.05). The density of ECL cells increased (65% vs 30%,P<0.01). CONCLUSIONS In DU and ZES,both the number and type of NE cells present some changes. Incresed gastrin in DU and ZES patients may be caused by the decrease of D cells and somatostatin secretion.

Liver fibrosis and hepatic stellate cells: Etiology, pathological hallmarks and therapeutic targets

Liver fibrosis is a reversible wound-healing process aimed at maintaining organ integrity, and presents as the critical pre-stage of liver cirrhosis, which will eventually progress to hepatocellular carcinoma in the absence of liver transplantation. Fibrosis generally results from chronic hepatic injury caused by various factors, mainly viral infection, schistosomiasis, and alcoholism; however, the exact pathological mechanisms are still unknown. Although numerous drugs have been shown to have antifibrotic activity in vitro and in animal models, none of these drugs have been shown to be efficacious in the clinic. Importantly, hepatic stellate cells(HSCs) play a key role in the initiation, progression, and regression of liver fibrosis by secreting fibrogenic factors that encourage portal fibrocytes, fibroblasts, and bone marrow-derived myofibroblasts to produce collagen and thereby propagate fibrosis. These cells are subject to intricate cross-talk with adjacent cells, resulting in scarring and subsequent liver damage. Thus, an understanding of the molecular mechanisms of liver fibrosis and their relationships with HSCs is essential for the discovery of new therapeutic targets. This comprehensive review outlines the role of HSCs in liver fibrosis and details novel strategies to suppress HSC activity, thereby providing new insights into potential treatments for liver fibrosis.

Differentiation potential of human adipose tissue derived stem cells into photoreceptors through explants culture and enzyme methods

AIM： To investigate the retinal photoreceptor differentiation potential of human orbital adipose tissue-derived stem cells （ADSCs） generated by enzyme （EN） and explant （EX） culture methods.METHODS： We investigated potentials of human orbital ADSCs to differentiate into photoreceptors through EN and EX culture methods. EN and EX orbital ADSCs were obtained from the same donor during rehabilitative orbital decompression, and then were subject to a 3-step induction using Noggin, DKK-1, IGF-1 and b-FGF at different time points for 38d. Stem cell, eye-field and photoreceptor-related gene and protein markers were measured by reverse transcription-polymerase chain reaction （RT-PCR） and immunofluorescent （IMF） staining.RESULTS： Both EX and EN orbital ADSCs expressed CD133, a marker of cell differentiation. Moreover, PAX6 and rhodopsin, markers of the retinal progenitor cells, were detected from EX and EN orbital ADSCs. In EX orbital ADSCs, PAX6 mRNA was detected on the 17th day and then the rhodopsin mRNA was detected on the 24th day. In contrast, the EN orbital ADSCs expressed PAX6 and rhodopsin mRNA on the 31st day. EX orbital ADSCs expressed rhodopsin protein on the 24th day, while EN orbital ADSCs expressed rhodopsin protein on the 31st day. CONCLUSION： Orbital ADSCs isolated by direct explants culture show earlier and stronger expressions of markers towards eye field and retinal photoreceptor differentiation than those generated by conventional EN method.

Activation of the TRAAK two-pore domain potassium channels in rd1 mice protects photoreceptor cells from apoptosis

AIM: To investigate the expression of TWIK-related arachidonic acid-stimulated K+ channel(TRAAK) in retinal degeneration mice(rd1) and further evaluate how TRAAK affect photoreceptor cell apoptosis.METHODS: The rd1 mice were distributed into blank(no treatment), control(1.4% DMSO, intraperitoneal injection) and riluzole groups(4 mg/kg·d, intraperitoneal injection) from postnatal 7 d to 10, 14 and 18 d;C57 group(no treatment), as age-matched wild-type control. The thickness of the outer nuclear layer(ONL) of retina was detected by paraffin section hematoxylin and eosin staining. The expression of TRAAK and the apoptosis of the ONL cells were detected by immunostaining, Western blotting, and real-time polymerase chain reaction. RESULTS: The channel agonist riluzole activated TRAAK and delayed the apoptosis of photoreceptor cells in ONL layer of rd1 mice. Both at mRNA and protein levels, after riluzole treatment, TRAAK expression was significantly upregulated, when compared with the control and blank group. Then we detected a series of apoptosis related mRNA and protein. The anti-apoptotic factor Bcl-2 downregulated and the pro-apoptotic factors Bax and cleaved-caspase-3 upregulated significantly. CONCLUSION: Riluzole elevates the expression of TRAAK and inhibits the development of apoptosis. Activation of TRAAK may have some potential effects to put off photoreceptor apoptosis.

Role of glycolysis in retinal vascular endothelium, glia, pigment epithelium, and photoreceptor cells and as therapeutic targets for related retinal diseases

Glycolysis produces large amounts of adenosine triphosphate(ATP)in a short time.The retinal vascular endothelium feeds itself primarily through aerobic glycolysis with less ATP.But when it generates new vessels,aerobic glycolysis provides rapid and abundant ATP support for angiogenesis,and thus inhibition of glycolysis in endothelial cells can be a target for the treatment of neovascularization.Aerobic glycolysis has a protective effect on Müller cells,and it can provide with a target for visual protection and maintenance of the blood-retinal barrier.Under physiological conditions,the mitochondria of RPE can use lactic acid produced by photoreceptor cells as an energy source to provide ATP for survival.In pathological conditions,because RPE cells avoid their oxidative damage by increasing glycolysis,a large number of glycolysis products accumulate,which in turn has a toxic effect on photoreceptor cells.This shows that stabilizing the function of RPE mitochondria may become a target for the treatment of diseases such as retinal degeneration.The decrease of aerobic glycolysis leads to the decline of photoreceptor cell function and impaired vision;therefore,aerobic glycolysis of stable photoreceptor cells provides a reliable target for delaying vision loss.It is of great significance to study the role of glycolysis in various retinal cells for the targeted treatment of ocular fundus diseases.

Reversing multiple age-related pathologies by controlling the senescence-associated secretory phenotype of stem cells

Regenerative medicine by cell transplantation is a novel therapy for treating end-stage organ failure and tissue damage. Cell-based therapy based on the transplantation of neural stem/progenitor cells （NSPCs） represents an attractive strategy for the treatment of neurodegenerative diseases, but obtaining large numbers of these cells is difficult and their differentiation potential is strictly restricted in a spatiotemporally-regulated manner during central nervous system （CNS） development. Therefore, embryonic stem cells and induced pluripotent stem cells represent an attractive alternative for cell-transplantation therapy in regenerative medicine.

Proteome changes during bone mesenchymal stem cell differentiation into photoreceptor-like cells in vitro

Human bone marrow stem cell (BMSC) may be directed to differentiate into multiple cell types, including adipocyte, chondrocyte, osteocyte and photoreceptor, among others. At present, little is known about the features of the BMSC and the protein control mechanism underlying their differentiation into photoreceptor-like cells. In the present study, BMSCs are induced to differentiate into photoreceptor-like cells in an in vitro model simulating the in vivo microenvironment. Up to 32 proteins are identified and differentially expressed through two-dimensional difference gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry to establish a differential protein database for photoreceptor-like cells from BMSC-induced differentiation. Western blot analysis further confirms the expression of some of the identified proteins. The present study proposes the total protein expression and possible molecular mechanism during the differentiation of BMSCs into photoreceptor cells.

Gastrointestinal B-cell lymphomas:From understanding B-cell physiology to classification and molecular pathology

The gut is the most common extranodal site where lymphomas arise. Although all histological lymphoma types may develop in the gut, small and large B-cell lymphomas predominate. The sometimes unexpected finding of a lymphoid lesion in an endoscopic biopsy of the gut may challenge both the clinician (who is not always familiar with lymphoma pathogenesis) and the pathologist (who will often be hampered in his/her diagnostic skill by the limited amount of available tissue). Moreover, the past 2 decades have spawned an avalanche of new data that encompasses both the function of the reactive B-cell as well as the pathogenic pathways that lead to its neoplastic counterpart, the B-cell lymphoma. Therefore, this review aims to offer clinicians an overview of B-cell lymphomas in the gut, and their pertinent molecular features that have led to new insights regarding lymphomagenesis. It addresses the question as how to incorporate all presently available information on normal and neoplastic B-cell differentiation, and how this knowledge can be applied in daily clinical practice (e.g., diagnostic tools, prognostic biomarkers or therapeutic targets) to optimalise the managment of this heterogeneous group of neoplasms.

Cytotoxic effect of specific T cells from mice with experimental autoimmune uveitis on murine photoreceptor cells

AIM:To investigate the cytotoxic effect of specific T cells from mice with experimental autoimmune uveitis(EAU)as well as their secreted interferon(IFN)-γand interleukin(IL)-17A on murine photoreceptor(661 W)cells.METHODS:An EAU model was established in female mice by injection of interphotoreceptor retinoid binding protein(IRBP)emulsion supplemented with complete Freund’s adjuvant(CFA)and Mycobacterium tuberculosis(TB).On day 12 after induction of EAU,specific T cells from spleen and lymph node tissues were isolated and cultured for 4 d and the levels of IFN-γand IL-17A in the supernatants were determined by enzyme-linked immunosorbent assays(ELISAs).T cells and their supernatants were added to 661 W cells to observe the alteration of cell morphology;IFN-γand IL-17A were separately added to 661 W cells to observe the effect of IFN-γand IL-17A on cell proliferation.RESULTS:The levels of IFN-γand IL-17A in the T cell supernatants were 1568.64±38.79 pg/m L and 1456.57±46.98 pg/mL,respectively.The supernatants apparently inhibited 661 W cell proliferation(P<0.05).T cells could also attach to the surface of 661 W cells,and IFN-γshowed a more serious cytotoxic effect on 661 W cells than IL-17A,inhibiting cell proliferation(P<0.01).CONCLUSION:IFN-γand IL-17A from T cells of EAU mice model can exert cytotoxic effects on murine photoreceptor cell proliferation,and IFN-γshows more serious cytotoxic effects on murine photoreceptor cells than IL-17A.

THE RELATIONSHIP BETWEEN SEROSAL TYPES, PATHOLOGICAL CHARACTERISTICS AND FREE CANCER CELLS IN THE PERITONEAL CAVITY OF GASTRIC CANCER PATIENTS

The relationship between free cancer cells and the pathological characteristics of gastric cancer were studied. Of 100 cases of gastric cancer, free cancer cells in the peritoneal cavity were detected in 32 cases (32%). Free cancer cells were most often found in tendonoid (62.2%) and colour diffused (60.0%) serosal types. When the area of serosal invasion was over 20 cm, the positive free cancer cell rate was 56.6% and only 2.5% if below 20 cm. Free cancer cells were related to the depth of cancer infiltration, often found in the S2 and S3 invasion layers, as well as to the pathological characteristics of gastric cancer. Free cancer cells were often seen in infiltrated type cancer, histologically poorly differentiated or undifferentiated adenocarcinoma. and nest or diffused growth types. In patients without peritoneal metastasis (P0), the positive rate was 26.1%. This study proved that Chen's serosal classification is correct and useful in assessing whether the cancer cells have penetrated through the serosa or not during surgery. Different treatments should be used in the cases with different serosal types. In addition to surgery, destruction of free cancer cells should be considered in tendonoid and colour diffused serosal types, so as to prevent peritoneal metastasis.

Human papillomavirus 16 E6 is associated with the nuclear matrix of esophageal carcinoma cells

AIM: To explore the etiologic role of HPV infection in esophageal carcinoma, and the association of HPV-16 E6with the nuclear metrix of carcinoma cells.METHODS: Two esophageal carcinoma cell lines, EC/CUHK1 and EC/CUHK2, were tested for HPV-16 E6subgenetic fragment by polymerase chain reaction amplification of virus DNA associated nuclear matrix. RT-PCR and immunocytochemistry were also used to visualizethe expression of E6 subgene in the cells.RESULTS: The HPV-16 E6 subgenetic fragment wes found to be present in nuclear metrix-associeted DNA, E6oncoprotein localized in the nucleus where it is tightly associated with nuclear matrix after sequential extraction in EC/CUHK2 cells. It was not detected, however, in EC/CUHK1 cells.CONCLUSION: The interaction between HPV-16 E6 and nuclear matrix may contribute to the virus induced carcinogenesis in esophageal carcinoma.

Osteoclastic and pleomorphic giant cell tumors of the pancreas:A review of clinical,endoscopic,and pathologic features

Giant cell tumors of the pancreas come in three varieties-osteoclastic,pleomorphic,and mixed histology.These tumors have distinctive endoscopic,clinical,and cytological features.Giant cell tumors have a controversial histogenesis,with some authors favoring an epithelial origin and others favoring a mesenchymal origin.The true origin of these lesions remains unclear at this time.These are also very rare tumors but proper identification and differentiation from more common pancreatic adenocarcinoma is important.The risk factors of these tumors and the prognosis may be different from those associated with standard pancreatic adenocarcinoma.Recognition of these differences can significantly affect patient care.These lesions have a unique appearance when imaged with endoscopic ultrasound(EUS),and these lesions can be diagnosed via EUS guided Fine Needle Aspiration(FNA).This manuscript will review the endoscopic,clinical,and pathologic features of these tumors.

Overexpressing NeuroD1 reprograms Müller cells into various types of retinal neurons

The onset of retinal degenerative disease is often associated with neuronal loss. Therefore, how to regenerate new neurons to restore vision is an important issue. NeuroD1 is a neural transcription factor with the ability to reprogram brain astrocytes into neurons in vivo. Here, we demonstrate that in adult mice, NeuroD1 can reprogram Müller cells, the principal glial cell type in the retina, to become retinal neurons. Most strikingly, ectopic expression of NeuroD1 using two different viral vectors converted Müller cells into different cell types. Specifically, AAV7 m8 GFAP681::GFP-ND1 converted Müller cells into inner retinal neurons, including amacrine cells and ganglion cells. In contrast, AAV9 GFAP104::ND1-GFP converted Müller cells into outer retinal neurons such as photoreceptors and horizontal cells, with higher conversion efficiency. Furthermore, we demonstrate that Müller cell conversion induced by AAV9 GFAP104::ND1-GFP displayed clear dose-and time-dependence. These results indicate that Müller cells in adult mice are highly plastic and can be reprogrammed into various subtypes of retinal neurons.

Intravitreal stem cell paracrine properties as a potential neuroprotective therapy for retinal photoreceptor neurodegenerative diseases

Retinal degenerations are the leading causes of irreversible visual loss worldwide. Many pathologies included under this umbrella involve progressive degeneration and ultimate loss of the photoreceptor cells, with age-related macular degeneration and inherited and ischemic retinal diseases the most relevant. These diseases greatly impact patients' daily lives, with accompanying marked social and economic consequences. However, the currently available treatments only delay the onset or slow progression of visual impairment, and there are no cures for these photoreceptor diseases. Therefore, new therapeutic strategies are being investigated, such as gene therapy, optogenetics, cell replacement, or cell-based neuroprotection. Specifically, stem cells can secrete neurotrophic, immunomodulatory, and anti-angiogenic factors that potentially protect and preserve retinal cells from neurodegeneration. Further, neuroprotection can be used in different types of retinal degenerative diseases and at different disease stages, unlike other potential therapies. This review summarizes stem cell-based paracrine neuroprotective strategies for photoreceptor degeneration, which are under study in clinical trials, and the latest preclinical studies. Effective retinal neuroprotection could be the next frontier in photoreceptor diseases, and the development of novel neuroprotective strategies will address the unmet therapeutic needs.

Cell transplantation to replace retinal ganglion cells faces challenges-the Switchboard Dilemma

The mammalian retina displays incomplete intrinsic regenerative capacities;therefore,retina degeneration is a major cause of irreversible blindness such as glaucoma,agerelated macular degeneration and diabetic retinopathy.These diseases lead to the loss of retinal cells and serious vision loss in the late stage.Stem cell transplantation is a great promising novel treatment for these incurable retinal degenerative diseases and represents an exciting area of regenerative neurotherapy.Several suitable stem cell sources for transplantation including human embryonic stem cells,induced pluripotent stem cells and adult stem cells have been identified as promising target populations.However,the retina is an elegant neuronal complex composed of various types of cells with different functions.The replacement of these different types of cells by transplantation should be addressed separately.So far,retinal pigment epithelium transplantation has achieved the most advanced stage of clinical trials,while transplantation of retinal neurons such as retinal ganglion cells and photoreceptors has been mostly studied in pre-clinical animal models.In this review,we opine on the key problems that need to be addressed before stem cells transplantation,especially for replacing injured retinal ganglion cells,may be used practically for treatment.A key problem we have called the Switchboard Dilemma is a major block to have functional retinal ganglion cell replacement.We use the public switchboard telephone network as an example to illustrate different difficulties for replacing damaged components in the retina that allow for visual signaling.Retinal ganglion cell transplantation is confronted by significant hurdles,because retinal ganglion cells receive signals from different interneurons,integrate and send signals to the correct targets of the visual system,which functions similar to the switchboard in a telephone network-therefore the Switchboard Dilemma.

Protective effects of naringenin eye drops on N-methylN-nitrosourea-induced photoreceptor cell death in rats

AIM:To investigate the effects of naringenin eye drops on N-methyl-N-nitrosourea (MNU)-induced photoreceptor cell death in rats.METHODS:Photoreceptor cell death was induced by single intraperitoneal injection of MNU(60 mg/kg)in rats.Both eyes of all animals were instilled with one drop of vehicle,0.5% or 1.0% naringenin eye drops three times per day from 7d before to 17d after MNU injection.Effects of naringenin on MNU-induced photoreceptor cell death were evaluated by electrophysiological and histological analysis.RESULTS:Flash electroretinography (FERG)and oscillatory potentials (OPs) recordings showed that the vehicle control group had remarkable reduction of amplitudes and prolongation of latency times.FERG and OPs responses were significantly reversed in MNUinduced rats treated with 0.5%or 1.0% naringenin eye drops compared with the vehicle control.The retinal morphological results showed that naringenin dosedependently preserved the outer nuclear layer,outer retina and total retina.CONCLUSION:These results indicate that topical treatment with naringenin eye drops prevented retinal neurons from MNU-induced structural and functional damages.

Stem cell therapy in ocular pathologies in the past 20 years

Stem cell therapies are successfully used in various fields of medicine.This new approach of research is also expanding in ophthalmology.Huge investments,resources and important clinical trials have been performed in stem cell research and in potential therapies.In recent years,great strides have been made in genetic research,which permitted and enhanced the differentiation of stem cells.Moreover,the possibility of exploiting stem cells from other districts(such as adipose,dental pulp,bone marrow stem cells,etc.)for the treatment of ophthalmic diseases,renders this topic fascinating.Furthermore,great strides have been made in biomedical engineering,which have proposed new materials and threedimensional structures useful for cell therapy of the eye.The encouraging results obtained on clinical trials conducted on animals have given a significant boost in the creation of study protocols also in humans.Results are limited to date,but clinical trials continue to evolve.Our attention is centered on the literature reported over the past 20 years,considering animal(the most represented in literature)and human clinical trials,which are limiting.The aim of our review is to present a brief overview of the main types of treatments based on stem cells in the field of ophthalmic pathologies.