Boron-peptide conjugates with angiopep-2 for boron neutron capture therapy.

Boron neutron capture therapy (BNCT) induces intracellular nuclear reaction to destroy cancer cells during thermal neutron irradiation. To selectively eliminate cancer cells but avoid harmful effects on normal tissues, novel boron-peptide conjugates with angiopep-2, namely ANG-B, were constructed and evaluated in preclinical settings. Boron-peptide conjugates were synthesized using solid-phase peptide synthesis, and the molecular mass was validated by mass spectrometry afterwards. Boron concentrations in 6 cancer cell lines and an intracranial glioma mouse model after treatments were analyzed by inductively coupled plasma atomic emission spectroscopy (ICP-AES). Phenylalanine (BPA) was tested in parallel for comparison. In vitro treatment with boron delivery peptides significantly increased boron uptake in cancer cells. BNCT with 5 mM ANG-B caused 86.5% ± 5.3% of clonogenic cell death, while BPA at the same concentration caused 73.3% ± 6.0% clonogenic cell death. The in vivo effect of ANG-B in an intracranial glioma mouse model was evaluated by PET/CT imaging at 31 days after BNCT. The mouse glioma tumours in the ANG-B-treated group were shrunk by 62.9% on average, while the BPA-treated tumours shrank by only 23.0%. Therefore, ANG-B is an efficient boron delivery agent, which has low cytotoxicity and high tumour-to-blood ratio. Based on these experimental results, we expected that ANG-B may leverage BNCT performance in clinical applications in future.

Human papillomavirus associated XPF deficiency increases alternative end joining and cisplatin sensitivity in head and neck squamous cell carcinoma.

OBJECTIVES: Human papillomavirus (HPV) positive head and neck squamous cell carcinoma (HNSCC) showed a considerably better prognosis with greater cisplatin sensitivity compared to their HPV-negative counterparts. Deciphering the underlying molecular mechanisms for HPV-induced cisplatin sensitivity is imperative to improve the prognosis of HPV-negative HNSCC. MATERIALS AND METHODS: The Fanconi anemia (FA) pathway status in HNSCC cells was analysed by detecting the cell cycle and chromosomal aberrations. XPF expression was validated using PCR, western blot, and immunohistochemistry. Droplet digital PCR and GFP expressing reporter assay were used to analyse the changes in alternative end-joining (alt-EJ) levels. The cisplatin sensitization was verified by cell proliferation assay, clonogenic cell survival assay, and TUNEL. RESULTS: HPV-positive HNSCC cells showed significant prolonged G2-M cell cycle arrest and aberrant chromosome formation under interstrand crosslinker treatment. Both mRNA and protein expression of XPF were considerably decreased in HPV-positive HNSCC, according to the analysis of cellular and clinical data. XPF inhibition upregulated the activity of the alt-EJ pathway in HPV-negative HNSCC cells by 32.02% (P < 0.001) but had little effect on HPV-positive HNSCC. Consistent with this, simultaneous suppression of XPF and alt-EJ enhanced cisplatin sensitivity of HPV-negative HNSCC in vitro and in vivo. CONCLUSION: HPV-positive HNSCC cells exhibit a profound FA pathway deficiency associated with reduced XPF expression. HNSCC cells with compromised XPF function are more reliant on the alt-EJ pathway for genomic stability. Combining FA and alt-EJ inhibition may be used to cope with the hard-to-treat HPV-negative HNSCC.

Detection of Alternative End-Joining in HNSC Cell Lines Using DNA Double-Strand Break Reporter Assays.

The main cellular pathways to repair DNA double-strand breaks (DSBs) and protect the integrity of the genome are homologous recombination (HR), non-homologous end-joining (NHEJ), and alternative end-joining (Alt-EJ). Polymerase theta-regulated Alt-EJ is an error-prone DSB repair pathway characterized by microhomology usage. Considering its importance in cancer treatment, technologies for detection of Alt-EJ in cancer cells may facilitate the study of the mechanisms of carcinogenesis and the development of new therapeutic targets. DSB reporter assay is the classical method for detecting Alt-EJ, which is primarily based on components of EJ2-puro cassette integration, I-SceI cleaving, and flow cytometry analysis. Here, we described an assay based on a modified I-Scel plasmid that can screen head and neck squamous cell carcinoma (HNSC) cells that were successfully transfected using selection medium with hygrovetine. We expect that this protocol will improve the fidelity and accuracy of reporter assays. Graphical abstract: Schematic overview of the workflow for establishment of Alt-EJ reporters.

CD44-fibrinogen binding promotes bleeding in acute promyelocytic leukemia by in situ fibrin(ogen) deposition.

Early hemorrhagic death is still the main obstacle for the successful treatment of acute promyelocytic leukemia (APL). However, the mechanisms underlying hemostatic perturbations in APL have not been fully elucidated. Here, we report that CD44 on the membrane of APL blasts and NB4 cells ligated bound fibrinogen, resulting in in situ deposition of fibrin and abnormal fibrin distribution. Clots formed by leukemic cells in response to CD44 and fibrinogen interaction exhibited low permeability and resistance to fibrinolysis. Using flow cytometry and confocal microscopy, we found that CD44 was also involved in platelet and leukemic cell adhesion. CD44 bound activated platelets but not resting platelets through interaction with P-selectin. APL cell-coated fibrinogen-activated platelets directly induce enhanced procoagulant activity of platelets. In vivo studies revealed that CD44 knockdown shortened bleeding time, increased the level of fibrinogen, and elevated the number of platelets by approximately twofold in an APL mouse model. Moreover, CD44 expression on leukemic cells in an APL mouse model was not only associated with bleeding complications but was also related to the wound-healing process and the survival time of APL mice. Collectively, our results suggest that CD44 may be a potential intervention target for preventing bleeding complications in APL.

CENPM upregulation by E5 oncoprotein of human papillomavirus promotes radiosensitivity in head and neck squamous cell carcinoma.

OBJECTIVES: This study aims to investigate how human papillomavirus (HPV) affects the key gene in the biological behaviors of head and neck squamous cell carcinoma (HNSCC) that leads to better response to radiotherapy. MATERIALS AND METHODS: The expression of key gene CENPM was analyzed using The Cancer Genome Atlas (TCGA) HNSCC data and HPV positive and HPV negative HNSCC tumors and cells. Assays with siRNAs, CRISPR/Cas9-based models, Western blot, qRT-PCR, ChIP, etc., were used to explore how HPV affects CENPM and response to radiotherapy for HNSCC. RESULTS: CENPM occupies the hub in the HPV-related gene network. HPV-positive HNSCC showed higher level of CENPM expression comparing with HPV-negative HNSCC. HPV E5 has the most pronounced impact on CENPM (R = 0.44, p = 0.00081). This might result from the binding of transcription factor E2F1 to CENPM. We further found that inhibition of CENPM expression in HPV-positive HNSCC cell line SCC47 increased resistance to X-ray radiation by approximately 59% under 2 Gy irradiation, which may be resulted from a reduced proportion of mitotic cells. CONCLUSION: HPV E5 enhances CENPM expression by transcription factor E2F1 in HNSCC, which results in a radiosensitive profile in cell cycle redistribution of HNSCC. Thus, HPV infection in HNSCC provides profound evidence that underscores the magnitude of E2F1 control of CENPM expression illustrating the potential clinical benefit of CENPM examination for difficult-to-treat HPV-negative cancers.

Extracellular Traps Increase Burden of Bleeding by Damaging Endothelial Cell in Acute Promyelocytic Leukaemia.

The rate of complete remission of acute promyelocytic leukemia (APL) is currently over 90% because of the use of all-trans retinoic acid (ATRA) with arsenic trioxide (ATO). However, hemorrhagic mortality has emerged as the most significant barrier to APL-induced remission. Neutrophils extracellular traps (NETs/ETs) cause vascular leakage by damaging the integrity of endothelial cells. We have previously demonstrated that APL cells treated with ATRA/ATO undergo a cell death process, releasing extracellular chromatin, termed ETosis/NETosis. However, the mechanism underlying the involvement of ETs in endothelial injury in APL remain largely unknown. Here, we analysed the ability of mature and immature neutrophils to release ETs, and their interaction with platelets (PLTs) in APL. Importantly, the effect of ETs on vascular endothelium in APL was discussed. Our results showed that the ability of immature neutrophils to release ETs was impaired in APL, whereas mature neutrophils produced ETs, which were associated with activated PLTs. Moreover, ATRA+ATO induced immature neutrophil differentiation, as well as increased the release of ETs from mature neutrophils. The excessive ETs damaged endothelial cells, causing blood cell leakage. Removing ETs using DNase 1 alleviated endothelial damage and improved blood cells leakage. Our results indicate that vascular endothelial injury is at least partially associated with ETs in APL, and that targeting ETs production may be an effective approach for relieving vascular leakage and reducing the burden of bleeding in APL.

Intestinal Damage in COVID-19: SARS-CoV-2 Infection and Intestinal Thrombosis.

The intestinal tract, with high expression of angiotensin-converting enzyme 2 (ACE2), is a major site of extrapulmonary infection in COVID-19. During pulmonary infection, the virus enters the bloodstream forming viremia, which infects and damages extrapulmonary organs. Uncontrolled viral infection induces cytokine storm and promotes a hypercoagulable state, leading to systemic microthrombi. Both viral infection and microthrombi can damage the gut-blood barrier, resulting in malabsorption, malnutrition, and intestinal flora entering the blood, ultimately increasing disease severity and mortality. Early prophylactic antithrombotic therapy can prevent these damages, thereby reducing mortality. In this review, we discuss the effects of SARS-CoV-2 infection and intestinal thrombosis on intestinal injury and disease severity, as well as corresponding treatment strategies.

Microvesicles, blood cells, and endothelial cells mediate phosphatidylserine-related prothrombotic state in patients with periodontitis.

BACKGROUND: Phosphatidylserine (PS) is essential for inflammation-associated thrombogenesis, but the exact effect of PS on the prothrombotic state in periodontitis is uncertain. This study aimed to determine the PS-related procoagulant state in patients with periodontitis. METHODS: A total of 138 patients with periodontitis were examined compared with 42 healthy controls. PS-exposing cells and microvesicles in blood samples were detected by confocal microscopy and flow cytometry. The clotting time assay and prothrombinase complex formation assay were used to measure the procoagulant activity of microvesicles, blood cells and endothelial cells. Periodontal clinical parameters and laboratory characteristics of patients with severe periodontitis were recorded and analyzed at baseline and 6 months after non-surgical periodontal therapy. RESULTS: Total PS-positive (PS+ ) microvesicles and the percentage of PS+ blood cells increased in patients with severe periodontitis compared with patients with moderate/mild periodontitis or healthy controls. Endothelial cells cultured in serum from patients with severe periodontitis expressed more PS compared with those cultured in serum from healthy controls. Specifically, PS exposure on blood cells and endothelial cells significantly decreased after inhibiting the effect of inflammatory cytokines. The elevated levels of PS+ cells and microvesicles in severe periodontitis shortened clotting time and led to increased prothrombinase complex formation. Non-surgical periodontal therapy significantly attenuated the release of microvesicles and the PS exposure of blood cells in severe periodontitis. CONCLUSIONS: The prothrombotic state of patients with periodontitis is mediated by PS+ cells and microvesicles stimulated by elevated levels of inflammatory cytokines.

Novel missense mutations of MVK and FDPS gene in Chinese patients with disseminated superficial actinic porokeratosis.

BACKGROUND AND AIMS: Porokeratosis (PK) is a heterogeneous group of cutaneous keratinization disorders and has five clinical subtypes. DSAP is the most common clinical subtype and is characterized by multiple small, annular, anhidrotic, keratotic lesions predominantly on sun-exposed areas of the skin. It is an autosomal dominantly inherited epidermal keratinization disorder. However, studies on its molecular basis is limited. MATERIALS AND METHODS: We performed mutation analysis of genes in four pedigrees and three sporadic cases of DSAP in the Chinese population. Genomic DNA was extracted from blood samples obtained from patients, unaffected family members, and 100 unrelated individuals. All exons and flanking intron sequences of the mevalonate kinase (MVK) and farnesyl diphosphate synthase (FDPS) genes were amplified. RESULTS: One missense mutation in exon 7 (C.G677A) of the MVK gene was identified in pedigree 3, and one missense mutation in exon 5 (C.C535T) of the FDPS gene was identified in sporadic case 3. No mutation was detected in the MVK and FDPS genes in the remaining three pedigrees and two sporadic cases with DSAP. CONCLUSION: Our results may be useful for genetic counseling and prenatal diagnosis of affected families and for expanding the repertoire of MVK and FDPS mutations underlying DSAP.

Effects of a School-Based Physical Activity Intervention for Obesity and Health-Related Physical Fitness in Adolescents With Intellectual Disability: Protocol for a Randomized Controlled Trial.

BACKGROUND: Childhood obesity accompanied by lower levels of health-related physical fitness (HRPF) is a major threat to public health both internationally and locally. Children with intellectual disability, especially adolescents, have a higher risk of being overweight/obese and having poor HRPF levels. Therefore, more interventions are needed to help this population attain their optimal health levels. However, there has been relatively limited research on this population compared with on their typically developing peers. OBJECTIVE: The proposed study aims to fill this knowledge gap by developing and examining the success of a physical activity (PA) intervention for the target population. METHODS: The proposed study will be a 12-week, school-based randomized controlled trial. The participants (N=48) will be recruited from special schools for students with mild intellectual disability and then randomly allocated to either the intervention group (IG) or the wait-list control group (CG). During the intervention period, the participants in the IG will receive a fun game-based moderate-to-vigorous PA (MVPA) training program (2 sessions/week, 60 minutes/session, for a total of 24 sessions). The intensity of the activities will increase in a progressive manner. Participants in the CG will receive no program during the study period, but the same PA program will be provided to them after the completion of the study. To observe and evaluate the sustaining effects of the intervention, follow-up testing will be scheduled for the participants 12 weeks after the intervention concludes. The study outcomes will include primary outcomes (obesity- and fitness-related outcomes) and a secondary outcome (blood pressure). All of the measurements will be taken at 3 time points. After the follow-up tests, the same PA training program will be provided to the participants in the CG. RESULTS: This study is ongoing. The participants were recruited from October 2020 to November 2020. The total duration of the study is 13 months. Study results are expected at the end of 2021. CONCLUSIONS: The proposed study is expected to reduce obesity and improve HRPF levels in children with intellectual disability. If proven effective, the intervention will be made accessible to more special schools and mainstream schools with students with intellectual disability. Furthermore, the study can serve as an example for international researchers, policy makers, and members of the public who are seeking to tackle the problem of obesity and poor HRPF among children with intellectual disability. TRIAL REGISTRATION: ClinicalTrials.gov NCT04554355; https://www.clinicaltrials.gov/ct2/show/NCT04554355. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): PRR1-10.2196/25838.

The Central Role of Extracellular Vesicles in the Mechanisms of Thrombosis in COVID-19 Patients With Cancer and Therapeutic Strategies.

Cancer patients have increased SARS-CoV-2 susceptibility and are prone to developing severe COVID-19 infections. The incidence of venous thrombosis is approximately 20% in COVID-19 patients with cancer. It has been suggested that thrombus formation has been suggested to correlate with severe clinical manifestations, mortality, and sequelae. In this review, we primarily elaborate on the pathophysiological mechanisms of thrombosis in COVID-19 patients with cancer, emphasize the role of microparticles (MPs) and phosphatidylserine (PS) in coagulation, and propose an antithrombotic strategy. The coagulation mechanisms of COVID-19 and cancer synergistically amplify the coagulation cascade, and collectively promotes pulmonary microvascular occlusion. During systemic coagulation, the virus activates immune cells to release abundant proinflammatory cytokines, referred to as cytokine storm, resulting in the apoptosis of tumor and blood cells and subsequent MPs release. Additionally, we highlight that tumor cells contribute to MPs and coagulation by apoptosis owing to insufficient blood supply. A positive feedback loop of cytokines storm and MPs storm promotes microvascular coagulation storm, leading to microthrombi formation and inadequate blood perfusion. Microthrombi-damaged endothelial cells (ECs), tumor, and blood cells further aggravate the apoptosis of the cells and facilitate MPs storm. PS, especially on MPs, plays a pivotal role in the blood coagulation process, contributing to clot initiation, amplification, and propagation. Since coagulation is a common pathway of COVID-19 and cancer, and associated with mortality, patients would benefit from antithrombotic therapy. The above results lead us to assert that early stage antithrombotic therapy is optimal. This strategy is likely to maintain blood flow patency contributing to viral clearance, attenuating the formation of cytokines and MPs storm, maintaining oxygen saturation, and avoiding the progress of the disease.

Neutrophil extracellular traps enhance procoagulant activity and thrombotic tendency in patients with obstructive jaundice.

BACKGROUND & AIMS: Patients with obstructive jaundice (OJ) are considered to be prothrombotic with increased risk of thromboembolism complications. The role of neutrophil extracellular traps (NETs) in procoagulant activity (PCA) and thrombosis risk in patients with OJ is unclear. In this study, we investigated NETs formation in OJ patients and the role of elevated unconjugated bilirubin (UCB) in inducing NETs, resulting in enhanced PCA and endothelial injury. METHODS: NETs of OJ patients and healthy controls were measured. NETs PCA was assessed via coagulation time (CT), fibrin formation and purified coagulation complex production assays. Visualization of NETs and mitochondrial reactive oxygen species (MitoROS) were performed with a fluorescence microscope. We further used confocal microscopy to quantify the exposure of phosphatidylserine (PS), fibrin strands and FVa/Xa on Human umbilical vein endothelial cells (HUVECs). RESULTS: Assessment of NETs components levels revealed greater NETs production in OJ patients than in healthy controls. Importantly, OJ-NETs were responsible for enhanced PCA. UCB induced NETs formation via MitoROS accumulation and mitochondrial mobilization. HUVECs cocultured with OJ NETs lost their cell-cell junctions and consequently converted to a procoagulant phenotype. The PCA was attenuated by using DNase I alone or in combination with lactadherin. CONCLUSIONS: Our results suggest that UCB-induced NETs play a prominent role in promoting the hypercoagulable and prothrombotic state in OJ patients. The increased MitoROS accumulation in neutrophils initiated NETosis. NETs are promising targets for indicating or improving coagulation disorders in OJ patients.

Phosphatidylserine-exposing blood cells, microparticles and neutrophil extracellular traps increase procoagulant activity in patients with pancreatic cancer.

Patients with pancreatic cancer (PC) are at increased risk of venous thrombosis, but the precise mechanisms of hypercoagulable state in PC remain unclear. We aimed to identify how phosphatidylserine positive (PS+) blood cells (BCs), PS+ microparticles (MPs) and neutrophil extracellular traps (NETs) regulate procoagulant activity (PCA) in PC, and to assess the relationship between PCA and PC staging. A total of 83 PC patients with different stages of disease were compared to 30 healthy controls, with confocal microscopy and flow cytometry used to assess MP and cellular PS exposure. MP and cell PCA was determined using both fibrin production assays and procoagulant enzyme complex analyses, and coagulation time was further measured. Patients with stage I PC and healthy controls exhibited significantly lower frequencies of PS+ MPs and BCs relative to those with more advanced disease, which may partly due to the increased levels of inflammation cytokines in advanced disease. Functional coagulation assays indicated that PS+ MPs and BCs derived from patients with stage II/III/IV PC directly contribute to elevated FXa, thrombin, and fibrin formation, and to more rapid coagulation relative to healthy control samples. In inhibition assays, lactadherin, which antagonizes PS, led to a roughly 80% inhibition of PCA. We further used isolated NETs to stimulate endothelial cells, revealing that this led to morphological changes including retraction from cell-cell junctions and a more pro-coagulative phenotype, with DNase I and activated protein C treatment reversing these changes. In patients with stage III PC, curative resection surgery significantly reduced PCA, whereas non-curative surgery did not have a marked impact based on studies of pre- and post-operative samples. These results highlight the pathogenic activity of PS+ cells, MPs, and NETs in promoting a prothrombotic environment within individuals suffering from advanced PC. Targeting PS and NETs in these patients may thus be a viable means of preventing pathological thrombosis.

Differentially expressed long non­coding RNAs and mRNAs in patients with IgA nephropathy.

Long non­coding RNAs (lncRNAs) have been reported to serve a crucial role in renal diseases; however, their role in immunoglobulin A nephropathy (IgAN) remains unclear. In the present study, peripheral blood mononuclear cells (PBMCs) were collected from both patients with IgAN and healthy controls. A microarray analysis was then performed to identify differentially expressed lncRNAs and mRNAs in PBMCs, which were confirmed by quantitative polymerase chain reaction. In addition, Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway and lncRNA­mRNA co­expression network analyses were conducted. The present study identified 167 differentially expressed lncRNAs and 94 differentially expressed mRNAs. Numerous GO terms, including innate immune response, inflammatory response, IPAF inflammasome complex and UDP­galactose:ß­N­acetylglucosamine ß­1, and 3­galactosyltransferase activity, were significantly enriched in the differentially expressed mRNAs. The top five KEGG signaling pathways included nucleotide­binding oligomerization domain­like receptor signaling pathway, hematopoietic cell lineage, inflammatory bowel disease, tumor necrosis factor signaling pathway and other types of O­glycan biosynthesis. In addition, a total of 149 lncRNAs were shown to interact with 7 mRNAs that were associated with the 'innate immune response' GO term. The results of the present study demonstrated that differentially expressed lncRNAs and mRNAs may have a role in the development of IgAN. These results may aid in the elucidation of a basic pathogenic mechanism, the identification of possible biomarkers and the generation of potential novel treatment strategies for IgAN.

Fenofibrate, a PPARα agonist, protect proximal tubular cells from albumin-bound fatty acids induced apoptosis via the activation of NF-kB.

Albumin-bound fatty acids is the main cause of renal damage, PPARα is responsible in the metabolism of fatty acids. Previous study found that PPARα played a protective role in fatty acids overload associated tubular injury. The aim of the present study is to investigate whether fenofibrate, a PPARα ligands, could contribute to the renoprotective action in fatty acids overload proximal tubule epithelial cells. We observed in HK-2 cells that fenofibrate significantly inhibited fatty acids bound albumin (FA-BSA) induced up-regulation of MCP-1 and IL-8. Treatment with fenofibrate attenuated renal oxidative stress induced by FA-BSA as evidenced by decreased MDA level, increased SOD activity and catalase, GPx-1 expression. FA-BSA induced apoptosis of HK-2 cells were also obviously prevented by fenofibrate. Furthermore, fenofibrate significantly increased the expression of PPARα mRNA and protein in FA-BSA treated cells. Finally, the activation of NF-kB induced by FA-BSA was markedly suppressed by fenofibrate. Taken together, our study describes a renoprotective role of fenofibrate in fatty acids associated tubular toxicity, and the transcriptional activation of PPARα and suppression of NF-kB were at least partially involved.

Vitiligo nonsurgical treatment: a review of latest treatment researches.

Vitiligo is a common depigmenting acquired disorder affecting about 1-2% of the world population, regardless of race, ethnic background, or gender. It is characterized by the appearance of milky white maculae because of a loss of melanocytes. The disfiguring nature of vitiligo causes high psychosocial morbidity. This is especially pronounced in populations with darker skin tone, likely because of the marked contrast. A variety of nonsurgical treatment regimens are currently employed in vitiligo. We reviewed the latest studies carried out on different nonsurgical treatment modalities used in vitiligo. All nonsurgical treatment aid to repigment or depigmentation the skin, however, many of them require a prolonged treatment course and may yield minimal results as well as carry unwanted side effects. There is a need for further research into the causes of vitiligo and into discovering better treatments.

Manganese-induced effects on cerebral trace element and nitric oxide of Hyline cocks.

Exposure to Manganese (Mn) is a common phenomenon due to its environmental pervasiveness. To investigate the Mn-induced toxicity on cerebral trace element levels and crucial nitric oxide parameters on brain of birds, 50-day-old male Hyline cocks were fed either a commercial diet or a Mn-supplemented diet containing 600, 900, 1,800 mg kg(-1). After being treated with Mn for 30, 60, and 90 days, the following were determined: the changes in contents of copper (Cu), iron (Fe), zinc (Zn), calcium (Ca), selenium (Se) in brain; inducible nitric oxide synthase-nitric oxide (iNOS-NO) system activity in brain; and histopathology and ultrastructure changes of cerebral cortex. The results showed that Mn was accumulated in brain and the content of Cu and Fe increased. However, the levels of Zn and Se decreased and the Ca content presented no obvious regularity. Exposure to Mn significantly elevated the content of NO and the expression of iNOS mRNA. Activity of total NO synthase (T NOS) and iNOS appeared with an increased tendency. These findings suggested that Mn exposure resulted in the imbalance of cerebral trace elements and influenced iNOS in the molecular level, which are possible underlying nervous system injury mechanisms induced by Mn exposure.

Protective effects of tubular liver-type fatty acid-binding protein against glomerular damage in murine IgA nephropathy.

BACKGROUND: Liver-type fatty acid-binding protein (L-FABP) in proximal tubules was reported to have renoprotective roles in experimental tubulointerstitial diseases via its anti-oxidative properties. Since tubuloglomerular cross-talk was recently discussed in the progression of renal diseases, to investigate whether tubular L-FABP may have an impact on the progression of glomerular damage, we induced IgA nephropathy (IgAN) in mice (Tg) transgenically tubular overexpressing human L-FABP (hL-FABP). METHODS: We reconstituted IgAN by bone marrow transplantation (BMT) from IgAN-prone mice into Tg and wild-type (WT) mice. Renal damage was evaluated at 6 and 12 weeks after BMT. During in vitro experiments, mesangial cells (MC) were stimulated by aggragated IgA (AIgA), and their supernatants (AIgA-MC medium) were collected. Stable cell line of mouse proximal tubular cell (mProx) transfected with or without hL-FABP gene was cultured with the AIgA-MC medium. RESULTS: Although mesangial IgA deposition and serum IgA level were not different between WT (WT/ddY) and Tg (Tg/ddY) recipients, WT/ddY mice showed a significantly higher urinary albumin level and mesangial matrix expansion with a significantly higher glomerular damage score. Furthermore, CD68 + macrophage infiltration was also significantly attenuated in Tg/ddY mice. Up-regulation of renal hL-FABP was associated with significant suppression of renal heme oxygenase-1 (HO-1) expression and accumulation of 4-hydroxy-2-nonenal (4-HNE) and MCP-1 expression in Tg/ddY mice. In vitro experiments showed that AIgA-MC medium and recombinant TNF-α significantly up-regulated hL-FABP expression, which was partially blocked by anti-TNF-α antibody, and major mediators of oxidative stress (HO-1 and 4-HNE) and inflammation (MCP-1). Importantly, such up-regulation of the mediators in mProx with hL-FABP was significantly suppressed much more than that in mProx. CONCLUSIONS: Tubular L-FABP activated by MC-origin humoral factors may lessen progression of glomerular damage at early stages of IgAN by reducing oxidative stress and inflammatory mediators.

The role of tumor necrosis factor-α converting enzyme in renal transplant rejection.

BACKGROUND: Tumor necrosis factor receptors (TNFR) are differentially regulated in human rejecting renal transplants. The TNF-α converting enzyme (TACE) regulates the membrane shedding of both receptors and TNF itself. We analyzed the expression and regulation of TACE in human rejecting renal transplants. METHODS: Samples from normal renal tissue or renal transplant undergoing severe acute rejection were immunostained for TACE using antibodies from different species. Human kidney epithelial cells were cultured and TACE plasmid was transfected to upregulate TACE expression. Cells were fractionated and immunoblotted for TACE, and ELISA was performed to test soluble TNFR2. RESULTS: We showed that TACE was upregulated mainly in tubular epithelial cells in acute rejecting kidney, where it colocalized with TNFR2. Epithelial cells with increased levels of TACE shed more soluble TNFR2 into culture media and even more after TACE activation by phorbol-12-myristate-13-acetate stimulation. The shedding could be completely blocked by the TACE inhibitor TNF-α protease inhibitor. CONCLUSION: Upregulation of TACE in epithelial cells in acute rejecting kidney could lead to more TNFR2 shedding and, hence, antagonize the proinflammatory effect of local TNF.