Therapeutic application of decellularized porcine small intestinal submucosa scaffold in conjunctiva reconstruction.

The objective of this study was to investigate the biological feasibility and surgical applicability of decellularized porcine small intestinal submucosa (DSIS) in conjunctiva reconstruction. A total of 52 Balb/c mice were included in the study. We obtained the DSIS by decellularization, evaluated the physical and biological properties of DSIS in vitro, and further evaluated the effect of surgical transplantation of DSIS scaffold in vivo. The histopathology and ultrastructural analysis results showed that the scaffold retained the integrity of the fibrous morphology while removing cells. Biomechanical analysis showed that the elongation at break of the DSIS (239.00 ± 12.51%) were better than that of natural mouse conjunctiva (170.70 ± 9.41%, P < 0.05). Moreover, in vivo experiments confirmed the excellent biocompatibility of the decellularized scaffolds. In the DSIS group, partial epithelialization occurred at day-3 after operation, and the conjunctival injury healed at day-7, which was significantly faster than that in human amniotic membrane (AM) and sham surgery (SHAM) group (P < 0.05). The number and distribution of goblet cells of transplanted DSIS were significantly better than those of the AM and SHAM groups. Consequently, the DSIS scaffold shows excellent biological characteristics and surgical applicability in the mouse conjunctival defect model, and DSIS is expected to be an alternative scaffold for conjunctival reconstruction.

Single-cell analysis reveals exosome-associated biomarkers for prognostic prediction and immunotherapy in lung adenocarcinoma.

BACKGROUND: Exosomes play a crucial role in tumor initiation and progression, yet the precise involvement of exosome-related genes (ERGs) in lung adenocarcinoma (LUAD) remains unclear. METHODS: We conducted a comprehensive investigation of ERGs within the tumor microenvironment (TME) of LUAD using single-cell RNA sequencing (scRNA-seq) analysis. Multiple scoring methods were employed to assess exosome activity (EA). Differences in cell communication were examined between high and low EA groups, utilizing the "CellChat" R package. Subsequently, we leveraged multiple bulk RNA-seq datasets to develop and validate exosome-associated signatures (EAS), enabling a multifaceted exploration of prognosis and immunotherapy outcomes between high- and low-risk groups. RESULTS: In the LUAD TME, epithelial cells demonstrated the highest EA, with even more elevated levels observed in advanced LUAD epithelial cells. The high-EA group exhibited enhanced intercellular interactions. EAS were established through the analysis of multiple bulk RNA-seq datasets. Patients in the high-risk group exhibited poorer overall survival (OS), reduced immune infiltration, and decreased expression of immune checkpoint genes. Finally, we experimentally validated the high expression of SEC61G in LUAD cell lines and demonstrated that knockdown of SEC61G reduced the proliferative capacity of LUAD cells using colony formation assays. CONCLUSION: The integration of single-cell and bulk RNA-seq analyses culminated in the development of the profound and significant EAS, which imparts invaluable insights for the clinical diagnosis and therapeutic management of LUAD patients.

Iron Homeostasis Disorder and Alzheimer's Disease.

Iron is an essential trace metal for almost all organisms, including human; however, oxidative stress can easily be caused when iron is in excess, producing toxicity to the human body due to its capability to be both an electron donor and an electron acceptor. Although there is a strict regulation mechanism for iron homeostasis in the human body and brain, it is usually inevitably disturbed by genetic and environmental factors, or disordered with aging, which leads to iron metabolism diseases, including many neurodegenerative diseases such as Alzheimer's disease (AD). AD is one of the most common degenerative diseases of the central nervous system (CNS) threatening human health. However, the precise pathogenesis of AD is still unclear, which seriously restricts the design of interventions and treatment drugs based on the pathogenesis of AD. Many studies have observed abnormal iron accumulation in different regions of the AD brain, resulting in cognitive, memory, motor and other nerve damages. Understanding the metabolic balance mechanism of iron in the brain is crucial for the treatment of AD, which would provide new cures for the disease. This paper reviews the recent progress in the relationship between iron and AD from the aspects of iron absorption in intestinal cells, storage and regulation of iron in cells and organs, especially for the regulation of iron homeostasis in the human brain and prospects the future directions for AD treatments.

Retracted: Morusinol Exhibits Selective and Potent Antitumor Activity Against Human Liver Carcinoma by Inducing Autophagy, G2/M Cell Cycle Arrest, Inhibition of Cell Invasion and Migration, and Targeting of Ras/MEK/ERK Pathway.

An editorial decision has been made to retract this manuscript due to breach of publishing guidelines, following the identification of non-original and manipulated figures. Reference: Zhen Zhu, Tianyu Xiao, Xuejiao Chang, Yanfei Hua, Jinli Gao: Morusinol Exhibits Selective and Potent Antitumor Activity Against Human Liver Carcinoma by Inducing Autophagy, G2/M Cell Cycle Arrest, Inhibition of Cell Invasion and Migration, and Targeting of Ras/MEK/ERK Pathway. Med Sci Monit 2019; 25:1864-1870. 10.12659/MSM.912992.

Primary malignant melanoma of the lung: a case report and literature review.

BACKGROUND: Malignant melanoma (MM) generally presents as a primary neoplasm of the skin, and most MM cases of the respiratory system are metastatic. Primary MM of the lung (PMML) is quite rare, and its diagnosis is relatively difficult. CASE PRESENTATION: We report the case of a 57-year-old male patient with PMML who denied any history of tumours. His initial complaint was frequent coughs with bloody sputum for 4 days. Chest radiography demonstrated a high-density shadow in the lower lobe of the right lung, which was suspected to be a large space-occupying lesion on subsequent computed tomography (CT) and to be a hypermetabolic tumour by positron emission tomography-CT. To confirm the diagnosis, exploratory surgery was performed. Finally, we confirmed the diagnosis of PMML. CONCLUSIONS: PMML is extremely rare and easily misdiagnosed as lung cancer. Because of its morphological and immunophenotypic variations, the diagnosis of PMML remains difficult. This case report discusses the diagnosis and case management of a patient while referring to the existing literature.

Morusinol Exhibits Selective and Potent Antitumor Activity Against Human Liver Carcinoma by Inducing Autophagy, G2/M Cell Cycle Arrest, Inhibition of Cell Invasion and Migration, and Targeting of Ras/MEK/ERK Pathway.

BACKGROUND Liver cancer is one of the most commonly diagnosed cancers across the globe. The treatment is often difficult as it is diagnosed mostly at advanced stages. Moreover, the lack efficacious and less toxic drugs are another problem in the treatment of liver cancer. Against this background, in this study we evaluated the anticancer activity of morusinol against SK-HEP-1 liver cancer cells. MATERIAL AND METHODS The proliferation rate of liver cancer cell line was investigated by MTT assay. Autophagy was detected by transmission electron microscopy and cell cycle analysis was performed by flow cytometry. The protein expression was examined by Western blotting. RESULTS Morusinol inhibited the proliferation of liver cancer SK-HEP-1 cells, with an IC50 of 20 µM against the SK-HEP-1liver cancer cells. Further investigations indicated that the antiproliferative effects of morusinol are due to initiation of autophagy and G2/M cell cycle arrest, which was also associated with altered expression of several important proteins. Morusinol also suppressed the migration and invasion of SK-HEP-1liver cancer cells, and it suppressed the expression of p-MEK and p-ERK, leading to suppression of the Raf/MEK/ERK signalling cascade. CONCLUSIONS We found that morusinol exerts significant anticancer and autophagic effects on liver cancer cells and our results suggest the potential of morusinol in treatment of liver cancer.

An Ns1abp-like gene promotes white spot syndrome virus infection by interacting with the viral envelope protein VP28 in red claw crayfish Cherax quadricarinatus.

Influenza A virus non-structural-1A binding protein (named as Ns1abp) was originally identified as a host protein from human that bound to the viral NS-1 protein. In our previous study, the expression of an Ns1abp-like gene (denoted as CqNs1abp-like gene) was found to be up-regulated in a transcriptome library from the haematopoietic tissue (Hpt) cells of red claw crayfish Cherax quadricarinatus post white spot syndrome virus (WSSV) infection. To elucidate the role of CqNs1abp-like gene involved in WSSV infection, we cloned the CqNs1abp-like gene in which the open reading frame was 2232 bp, encoding 743 amino acids with two typical domains of one BTB (Broad-Complex, Tramtrack and Bric a brac) domain at N-terminal and six Kelch domains at C-terminal. The gene expression profile showed that the mRNA transcript of CqNs1abp-like gene was widely expressed in all the tested tissues with highest expression in nerve, relatively high expression in Hpt and lowest expression in eyestalk. Importantly, both the WSSV entry and the viral replication were significantly reduced in Hpt cells after gene silencing of CqNs1abp-like gene. By using protein pull-down assay, we found that the recombinant BTB domain, six Kelch domains and CqNs1abp-like intact protein were all bound to the WSSV envelope protein VP28, respectively, in which the BTB domain showed slightly less binding affinity than that of the six Kelch domains or the recombinant intact protein. Besides, the WSSV entry into Hpt cells was clearly decreased when the virus was pre-incubated with the recombinant BTB domain, six Kelch domains, or the recombinant CqNs1abp-like intact protein, respectively, suggesting that the CqNs1abp-like gene was likely to function as a putative recognition molecular towards WSSV infection in a crustacean C. quadricarinatus. Taken together, these data shed new light on the mechanism of WSSV infection and a putatively novel target on anti-WSSV infection in crustacean farming.

A thymosin repeated protein1 reduces white spot syndrome virus replication in red claw crayfish Cherax quadricarinatus.

The ß-thymosins are a group of structurally related, highly conserved intracellular small peptides in vertebrates with various biological functions, including cytoskeletal remodeling, neuronal development, cell migration, cell survival, tissue repair and inhibition of inflammation. In contrast to vertebrates, the function of ß-thymosin is not fully understood in crustaceans. Previously, we found that a thymosin-repeated protein1 (CqTRP1) gene was up-regulated after white spot syndrome virus (WSSV) challenge in hematopoietic tissue (Hpt) cells from the red claw crayfish Cherax quadricarinatus. To further identify the effect of CqTRP1 on WSSV infection, a full length cDNA sequence of ß-thymosin homologue was cloned and analyzed from red claw crayfish followed by functional study. The CqTRP1 cDNA contains an open reading frame of 387 nucleotides encoding a protein of 129 amino acids with a putative molecular mass of 14.3 kDa. The amino acid sequence showed high identity with other ß-thymosins and contained three characteristic thymosin ß actin-binding motifs, suggesting that CqTRP1 was a member of the ß-thymosin family. Tissue distribution analysis revealed a ubiquitous presence of CqTRP1 in all the examined tissues with the highest expression in hemocytes, Hpt and gonad at the transcriptional level. Interestingly, the gene silencing of endogenous CqTRP1 by RNAi enhanced the WSSV replication in Hpt cells. Meanwhile, the WSSV replication was significantly reduced in the Hpt cell cultures if overloaded with a recombinant CqTRP1. Taken together, these data clearly indicated that CqTRP1 was likely to be associated with the anti-WSSV response in a crustacean C. quadricarinatus, which provides new strategy against white spot disease in crustacean aquaculture.

A CqFerritin protein inhibits white spot syndrome virus infection via regulating iron ions in red claw crayfish Cherax quadricarinatus.

It is well known that iron is an essential element for all living organism. The intracellular iron availability is also important for the host's innate immune response to various pathogens, in which the iron homeostasis can be regulated by ferritin due to its iron storage property. In this study, a full-length cDNA sequence of ferritin (named as CqFerritin) was identified with 1410 bp from red claw crayfish Cherax quadricarinatus, which contained an open reading frame of 513 bp, encoding 170 amino acids with a conserved ferritin domain. Tissue distribution analysis demonstrated that CqFerritin was widely expressed in various tissues with high presence in haemocyte, haematopoietic tissue (Hpt) and heart, while lowest expression in hepatopancreas. In addition, loss-of-function of CqFerritin by gene silencing resulted in significantly higher expression of an envelope protein VP28 of white spot syndrome virus (WSSV) in red claw crayfish Hpt cell cultures, indicating the potential antiviral response of CqFerritin. To further explore the effect on WSSV replication by CqFerritin, recombinant CqFerritin protein (rCqFerritin) was transfected into Hpt cells followed by WSSV infection. Importantly, the replication of WSSV was obviously decreased in Hpt cells if transfected with rCqFerritin protein, suggesting that CqFerritin had clearly negative effect on WSSV infection. Furthermore, intracellular accumulation of iron ions was found to promote the WSSV replication in a dose-dependent manner, illustrating that the iron level regulated by CqFerritin was likely to be vital for WSSV infection in red claw crayfish. Taken together, these data suggest that CqFerritin plays an important role in immune defense against WSSV infection in a crustacean C. quadricarinatus.

Differential protein expression using proteomics from a crustacean brine shrimp (Artemia sinica) under CO2-driven seawater acidification.

Gradually increasing atmospheric CO2 partial pressure (pCO2) has caused an imbalance in carbonate chemistry and resulted in decreased seawater pH in marine ecosystems, termed seawater acidification. Anthropogenic seawater acidification is postulated to affect the physiology of many marine calcifying organisms. To understand the possible effects of seawater acidification on the proteomic responses of a marine crustacean brine shrimp (Artemia sinica) three groups of cysts were hatched and further raised in seawater at different pH levels (8.2 as control and 7.8 and 7.6 as acidification stress levels according to the predicted levels at the end of this century and next century, respectively) for 1, 7 and 14 days followed by examination of the protein expression changes via two-dimensional gel electrophoresis. Searches of protein databases revealed that 67 differential protein spots were altered due to lower pH level (7.6 and 7.8) stress in comparison to control groups (pH 8.2) by mass spectrometry. Generally, these differentially expressed proteins included the following: 1) metabolic process-related proteins involved in glycolysis and glucogenesis, nucleotide/amino acid/fatty acid metabolism, protein biosynthesis, DNA replication and apoptosis; 2) stress response-related proteins, such as peroxiredoxin, thioredoxin peroxidase, 70-kDa heat shock protein, Na/K ATPase, and ubiquinol-cytochrome c reductase; 3) immune defence-related proteins, such as prophenoloxidase and ferritin; 4) cytoskeletal-related proteins, such as myosin light chain, TCP1 subunit 2, tropomyosin and tubulin alpha chain; and 5) signal transduction-related proteins, such as phospholipase C-like protein, 14-3-3 zeta, translationally controlled tumour protein and RNA binding motif protein. Taken together, these data support the idea that CO2-driven seawater acidification may affect protein expression in the crustacean A. sinica and possibly also in other species that feed on brine shrimp in the ecosystem, particularly marine food webs.

White spot syndrome virus entry is dependent on multiple endocytic routes and strongly facilitated by Cq-GABARAP in a CME-dependent manner.

White spot syndrome virus (WSSV) is a lethal pathogen of shrimp and many other crustaceans, including crayfish. However, the molecular mechanism underlying its cellular entry remains elusive due to the lack of shrimp cell lines for viral propagation. Crayfish hematopoietic tissue (Hpt) cell culture was recently established as a good model for WSSV infection study. Here, we showed that multiple endocytic routes, including clathrin-mediated endocytosis (CME), macropinocytosis and caveolae-mediated endocytosis, were indispensably employed for the viral entry into Hpt cell of the crayfish Cherax quadricarinatus. Intriguingly, cellular autophagic activity was positively correlated with efficient viral entry, in which a key autophagy-related protein, γ-aminobutyric acid receptor-associated protein (Cq-GABARAP), that not only localized but also co-localized with WSSV on the Hpt cell membrane, strongly facilitated WSSV entry by binding to the viral envelope VP28 in a CME-dependent manner that was negatively regulated by Cq-Rac1. Furthermore, cytoskeletal components, including Cq-ß-tubulin and Cq-ß-actin, bound to both recombinant rCq-GABARAP and WSSV envelope proteins, which likely led to viral entry promotion via cooperation with rCq-GABARAP. Even under conditions that promoted viral entry, rCq-GABARAP significantly reduced viral replication at an early stage of infection, which was probably caused by the formation of WSSV aggregates in the cytoplasm.

Diagnosis and treatment of autoimmune pancreatitis: experience with 100 patients.

BACKGROUND: Autoimmune pancreatitis (AIP) is increasingly recognized as a unique subtype of pancreatitis. This study aimed to analyze the diagnosis and treatment of AIP patients from a tertiary care center in China. METHODS: One hundred patients with AIP who had been treated from January 2005 to December 2012 in our hospital were enrolled in this study. We retrospectively reviewed the data of clinical manifestations, laboratory tests, imaging examinations, pathological examinations, treatment and outcomes of the patients. RESULTS: The median age of the patients at onset was 57 years (range 23-82) with a male to female ratio of 8.1:1. The common manifestations of the patients included obstructive jaundice (49 patients, 49.0%), abdominal pain (30, 30.0%), and acute pancreatitis (11, 11.0%). Biliary involvement was one of the most extrapancreatic manifestations (64, 64.0%). Fifty-six (56.0%) and 43 (43.0%) patients were classified into focal-type and diffuse-type respectively according to the imaging examinations. The levels of serum IgG and IgG4 were elevated in 69.4% (43/62) and 92.0% (69/75) patients. Pathological analysis of specimens from 27 patients supported the diagnosis of lymphoplasmacytic sclerosing pancreatitis, and marked (>10 cells/HPF) IgG4 positive cells were found in 20 (74.1%) patients. Steroid treatment and surgery as the main initial treatments were given to 41 (41.0%) and 28 (28.0%) patients, respectively. The remission rate after the initial treatment was 85.0%. Steroid was given as the treatment after relapse in most of the patients and the total remission rate at the end of follow-up was 96.0%. CONCLUSIONS: Clinical manifestations, laboratory tests, imaging and pathology examinations in combination could increase the diagnostic accuracy of AIP. Steroid treatment with an initial dose of 30 or 40 mg prednisone is effective and safe in most patients with AIP.

Upregulation of Wnt5a promotes epithelial-to-mesenchymal transition and metastasis of pancreatic cancer cells.

BACKGROUND: Pancreatic cancer is one of the most lethal cancers worldwide. The aim of this study was to determine the expression pattern, clinical significance, and biological functions of Wnt5a in pancreatic cancer. METHODS: Immunohistochemistry was performed to examine Wnt5a expression in 134 surgically resected pancreatic adenocarcinoma and adjacent normal pancreatic tissues. Associations of Wnt5a expression with clinicopathological factors and cancer-specific survival were analyzed. The effects of Wnt5a overexpression or silencing on the invasiveness and epithelial-to-mesenchymal transition (EMT) of pancreatic cancer cells were studied. Silencing of ß-catenin by small interfering RNA was done to determine its role in the Wnt5a-mediated tumor phenotype. RESULTS: The percentage of Wnt5a positive expression showed a bell-shaped pattern in pancreatic cancer tissues, peaking in well-differentiated carcinomas. The median cancer-specific survival was comparable between patients with positive versus negative expression of Wnt5a. Overexpression of Wnt5a promoted the migration and invasion of pancreatic cancer cells, whereas Wnt5a depletion had an inhibitory effect. In an orthotopic pancreatic cancer mouse model, Wnt5a overexpression resulted in increased invasiveness and metastasis, coupled with induction of EMT in tumor cells. Treatment with recombinant Wnt5a elevated the nuclear ß-catenin level in pancreatic cancer cells, without altering the Ror2 expression. Targeted reduction of ß-catenin antagonized exogenous Wnt5a-induced EMT and invasiveness in pancreatic cancer cells. CONCLUSION: Upregulation of Wnt5a promotes EMT and metastasis in pancreatic cancer models, which involves activation of ß-catenin-dependent canonical Wnt signaling. These findings warrant further investigation of the clinical relevance of Wnt5 upregulation in pancreatic cancer.

[Clinicopathologic characteristics of fibrous mass-forming chronic pancreatitis].

OBJECTIVE: To investigate clinicopathological features of fibrous mass-forming chronic pancreatitis (FMCP), to compare clinicopathological and immunohistochemical characteristics between autoimmune pancreatitis (AIP) and fibrous mass-forming non-autoimmune pancreatitis (nAIP) and to provide evidence for pathological diagnosis, differential diagnosis and clinical treatment strategy. METHODS: Clinicopathological features were analyzed in 81 cases of FMCP. Infiltrating IgG4(+) plasmacytes were counted by immunohistochemical staining. RESULTS: Among 81 cases of FMCP, 20 cases were diagnosed as AIP and 61 cases were interpreted as nAIP. AIP was more common in males over 50 years, whereas nAIP was seen in much younger patients (P = 0.001). The amount of inflammatory cells in the stroma of AIPs was remarkable higher than that in nAIPs (P = 0.002). The incidence of neuritis in AIPs (100%, 20/20) was also higher compared with that of nAIPs (75.4%, 46/61; P = 0.017). Storiformed-fibrosis was more common in AIPs (95.0%, 19/20) than in nAIPs (1.6%, 1/61;P = 0.000). Pancreatic intraepithelial neoplasia (PanIN) was observed in 50.0%(10/20) of AIPs and 32.8%(20/61) of nAIPs, with a greater severity observed in AIPs (P = 0.031). Tubular complex (TC) was more commonly observed in AIPs (65.0%, 13/20) than nAIPs (26.2%, 16/61;P = 0.002). Among 81 cases of FMCP, 61 cases had less than 11 IgG4(+) plasmacytes /HPF, 7 cases had 10-30/HPF and 13 cases had over 30/HPF. CONCLUSIONS: FMCPs include both AIP and nAIP. AIP has distinct pathological features and the presence of IgG4(+) plasmacyte is an important diagnostic parameter. FMCP appears to be an important precancerous lesion of pancreatic ductal adenocarcinoma. Surgery may be considered for patients with FMCP due to its mass-forming nature. In contrast, patients with AIP are treated medically due to its steroid-responsiveness. Therefore, accurate and timely diagnosis of AIP is of clinical relevance to avoid unnecessary surgical complications and to prevent progression of the disease.