Neural Precursor-Derived Pleiotrophin Mediates Subventricular Zone Invasion by Glioma.

The lateral ventricle subventricular zone (SVZ) is a frequent and consequential site of pediatric and adult glioma spread, but the cellular and molecular mechanisms mediating this are poorly understood. We demonstrate that neural precursor cell (NPC):glioma cell communication underpins this propensity of glioma to colonize the SVZ through secretion of chemoattractant signals toward which glioma cells home. Biochemical, proteomic, and functional analyses of SVZ NPC-secreted factors revealed the neurite outgrowth-promoting factor pleiotrophin, along with required binding partners SPARC/SPARCL1 and HSP90B, as key mediators of this chemoattractant effect. Pleiotrophin expression is strongly enriched in the SVZ, and pleiotrophin knock down starkly reduced glioma invasion of the SVZ in the murine brain. Pleiotrophin, in complex with the binding partners, activated glioma Rho/ROCK signaling, and ROCK inhibition decreased invasion toward SVZ NPC-secreted factors. These findings demonstrate a pathogenic role for NPC:glioma interactions and potential therapeutic targets to limit glioma invasion. PAPERCLIP.

The matricellular protein SPARC induces inflammatory interferon-response in macrophages during aging.

The risk of chronic diseases caused by aging is reduced by caloric restriction (CR)-induced immunometabolic adaptation. Here, we found that the matricellular protein, secreted protein acidic and rich in cysteine (SPARC), was inhibited by 2 years of 14% sustained CR in humans and elevated by obesity. SPARC converted anti-inflammatory macrophages into a pro-inflammatory phenotype with induction of interferon-stimulated gene (ISG) expression via the transcription factors IRF3/7. Mechanistically, SPARC-induced ISGs were dependent on toll-like receptor-4 (TLR4)-mediated TBK1, IRF3, IFN-ß, and STAT1 signaling without engaging the Myd88 pathway. Metabolically, SPARC dampened mitochondrial respiration, and inhibition of glycolysis abrogated ISG induction by SPARC in macrophages. Furthermore, the N-terminal acidic domain of SPARC was required for ISG induction, while adipocyte-specific deletion of SPARC reduced inflammation and extended health span during aging. Collectively, SPARC, a CR-mimetic adipokine, is an immunometabolic checkpoint of inflammation and interferon response that may be targeted to delay age-related metabolic and functional decline.

Molecular divergence of mammalian astrocyte progenitor cells at early gliogenesis.

During mammalian brain development, how different astrocytes are specified from progenitor cells is not well understood. In particular, whether astrocyte progenitor cells (APCs) start as a relatively homogenous population or whether there is early heterogeneity remains unclear. Here, we have dissected subpopulations of embryonic mouse forebrain progenitors using single-cell transcriptome analyses. Our sequencing data revealed two molecularly distinct APC subgroups at the start of gliogenesis from both dorsal and ventral forebrains. The two APC subgroups were marked, respectively, by specific expression of Sparc and Sparcl1, which are known to function in mature astrocytes with opposing activities for regulating synapse formation. Expression analyses showed that SPARC and SPARCL1 mark APC subgroups that display distinct temporal and spatial patterns, correlating with major waves of astrogliogenesis during development. Our results uncover an early molecular divergence of APCs in the mammalian brain and provide a useful transcriptome resource for the study of glial cell specification.

Peritumoral SPARC expression induced by exosomes from nasopharyngeal carcinoma infected Epstein-Barr virus: A poor prognostic marker.

Epstein-Barr virus (EBV)-associated nasopharyngeal carcinoma (NPC) cells have high metastatic potential. Recent research has revealed that the interaction of between tumor cells and the surrounding stroma plays an important role in tumor invasion and metastasis. In this study, we showed the prognostic value of expression of SPARC, an extracellular matrix protein with multiple cellular functions, in normal adjacent tissues (NAT) surrounding NPC. In the immunohistochemical analysis of 51 NPC biopsy specimens, SPARC expression levels were significantly elevated in the NAT of EBER (EBV-encoded small RNA)-positive NPC compared to that in the NAT of EBER-negative NPC. Moreover, increased SPARC expression in NAT was associated with a worsening of overall survival. The enrichment analysis of RNA-seq of publicly available NPC and NAT surrounding NPC data showed that high SPARC expression in NPC was associated with epithelial mesenchymal transition promotion, and there was a dynamic change in the gene expression profile associated with interference of cellular proliferation in NAT, including SPARC expression. Furthermore, EBV-positive NPC cells induce SPARC expression in normal nasopharyngeal cells via exosomes. Induction of SPARC in cancer-surrounding NAT cells reduced intercellular adhesion in normal nasopharyngeal structures and promoted cell competition between cancer cells and normal epithelial cells. These results suggest that epithelial cells loosen their own binding with the extracellular matrix as well as stromal cells, facilitating the invasion of tumor cells into the adjacent stroma by activating cell competition. Our findings reveal a new mechanism by which EBV creates a pro-metastatic microenvironment by upregulating SPARC expression in NPC.

Exploring the roles of SPARC as a proinflammatory factor and its potential as a novel therapeutic target against cardiovascular disease.

Cardiovascular disease (CVD) is a leading cause of death worldwide, and the number of CVD patients continues to increase despite extensive research and developments in this field. Chronic inflammation is a pivotal pathological component of CVD, and unveiling new proinflammatory factors will help devise novel preventive and therapeutic strategies. The extracellular matrix (ECM) not only provides structural support between cells, but also contributes to cellular functions. Secreted protein acidic and rich in cysteine (SPARC) is a collagen-binding matricellular protein that is particularly induced during development and tissue remodeling. A proinflammatory role for SPARC has been demonstrated in various animal models, such as in the lipopolysaccharide-induced footpad model and dextran sodium sulfate-induced colitis model. Recent clinical studies reported a positive correlation between elevated plasma SPARC levels and hypertension, obesity, and the inflammatory marker high-sensitive C-reactive protein. In addition, SPARC gene deletion attenuates the cardiac injury induced by aging, myocardial infarction, and pressure-load, suggesting that SPARC has deleterious effects on CVD. This review summarizes the regulatory and proinflammatory mechanisms of SPARC on CVD, chronic kidney disease (CKD) and cerebrovascular disease, and discusses the rationale behind measuring SPARC as a biomarker of CVD, and the effects of inhibition of SPARC in the prevention and treatment of CVD.

Effect of Sparc knockout on the extracellular matrix of mouse periodontal ligament cells.

The periodontal ligament (PDL) is a critical component in maintaining tooth stability. It is composed of cells and an extracellular matrix (ECM), each with unique roles in tissue function and homeostasis. Secreted protein acidic and rich in cysteine (SPARC), a calcium-binding matricellular glycoprotein, plays a crucial role in regulating ECM assembly and turnover, alongside facilitating cellular-ECM interactions. In the present study, mass spectrometry-based proteomics was used to assess the impacts of Sparc-knockout (KO) on PDL-derived cells. Results demonstrated that Sparc-KO significantly reduces ECM production and alters its composition with increased levels of type I collagen. Despite this increase in Sparc-KO, type I collagen was not likely to be effectively integrated into the fibrils due to collagen cross-linking impairment. Furthermore, the pathway and process enrichment analyses suggested that SPARC plays a protective role against ECM degradation by antagonistically interacting with cell-surface collagen receptors. These findings provide detailed insights into the multifaceted role of SPARC in ECM organization, including its impact on ECM production, collagen regulation, and interactions with various cellular compartments. A better understanding of these complex mechanisms is crucial for comprehending the causes of periodontal disease and tissue regeneration, where precise control of ECM organization is necessary.

SPARC-YAP/TAZ inhibition prevents the fibroblasts-myofibroblasts transformation.

BACKGROUND: Fibrotic scar is a severe side effect of trabeculectomy, resulting in unsatisfactory outcomes for glaucoma surgery. Accumulating evidence showed human Tenon's fibroblasts (HTFs) play an important role in fibrosis formation. We previously reported that the aqueous level of secreted protein acidic and rich in cysteine (SPARC) was higher in the patients with primary angle closure glaucoma, which was associated with the failure of trabeculectomy. In this study, the potential effect and mechanism of SPARC in promoting fibrosis were explored by using HTFs. METHODS: HTFs were employed in this study and examined under a phase-contrast microscope. Cell viability was determined by CCK-8. The expressions of SPARC-YAP/TAZ signaling and the fibrosis-related markers were examined with reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR), Western blot, and immunofluorescence, subcellular fractionation was conducted to further determined the variation of YAP and phosphorylated YAP. The differential gene expressions were analyzed with RNA sequencing (RNAseq), followed by Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses. RESULTS: Exogenous SPARC induced HTFs-myofibroblast transformation, as evidenced by the increased expression of α-SMA, collagen I and fibronectin in both protein and mRNA levels. SPARC knockdown decreased the expressions of the above genes in TGF-ß2-treated HTFs. KEGG analysis showed that the Hippo signaling pathway was mostly enriched. SPARC treatment increased the expressions of YAP, TAZ, CTGF and CYR61 as well as enhanced YAP translocation from cytoplasm to nucleus, and decreased the phosphorylation of YAP and LAST1/2, which was reversed by SPARC knockdown. Knockdown of YAP1 decreased the fibrosis-related markers, such as α-SMA, collagen I and Fibronectin, in SPARC-treated HTFs. CONCLUSIONS: SPARC induced HTFs-myofibroblast transformation via activating YAP/TAZ signaling. Targeting SPARC-YAP/TAZ axis in HTFs might provide a novel strategy for inhibiting fibrosis formation after trabeculectomy.

FGF-2-dependent signaling activated in aged human skeletal muscle promotes intramuscular adipogenesis.

Aged skeletal muscle is markedly affected by fatty muscle infiltration, and strategies to reduce the occurrence of intramuscular adipocytes are urgently needed. Here, we show that fibroblast growth factor-2 (FGF-2) not only stimulates muscle growth but also promotes intramuscular adipogenesis. Using multiple screening assays upstream and downstream of microRNA (miR)-29a signaling, we located the secreted protein and adipogenic inhibitor SPARC to an FGF-2 signaling pathway that is conserved between skeletal muscle cells from mice and humans and that is activated in skeletal muscle of aged mice and humans. FGF-2 induces the miR-29a/SPARC axis through transcriptional activation of FRA-1, which binds and activates an evolutionary conserved AP-1 site element proximal in the miR-29a promoter. Genetic deletions in muscle cells and adeno-associated virus-mediated overexpression of FGF-2 or SPARC in mouse skeletal muscle revealed that this axis regulates differentiation of fibro/adipogenic progenitors in vitro and intramuscular adipose tissue (IMAT) formation in vivo. Skeletal muscle from human donors aged >75 y versus <55 y showed activation of FGF-2-dependent signaling and increased IMAT. Thus, our data highlights a disparate role of FGF-2 in adult skeletal muscle and reveals a pathway to combat fat accumulation in aged human skeletal muscle.

Interference with SPARC inhibits Benzophenone-3 induced ferroptosis in osteoarthritis: Evidence from bioinformatics analyses and biological experimentation.

The aim of this study is to conduct a thorough evaluation of the association between Benzophenone-3 (BP-3) exposure and OA, offering critical insights into the underlying mechanisms involved. The National Health and Nutrition Examination Survey (NHANES) database was utilized to investigate the correlation between BP-3 and osteoarthritis. Proteomic sequencing from clinical sample and the PharmMapper online tool were employed to predict the biological target of BP-3. Cellular molecular assays and transfection studies were performed to verify the prediction from bioinformatics analyses. Through cross-sectional analysis of the NHANES database, we identified BP-3 as a risk factor for OA development. The results of proteomic sequencing showed that Secreted Protein Acidic and Rich in Cysteine (SPARC) was significantly elevated in the area of damage compared to the undamaged area. SPARC was also among the potential biological targets of BP-3 predicted by the online program. Through in vitro cell experiments, we further determined that the toxicological effects of BP-3 may be due to SPARC, which elevates intracellular GPX4 levels, activates the glutathione system, and promotes lipid peroxidation to mitigate ferroptosis. Inhibiting SPARC expression has been shown to reduce inflammation and ferroptosis in OA contexts. This research provides an expansive understanding of BP-3's influence on osteoarthritis development. We have identified SPARC as a potent target for combating chondrocyte ferroptosis in BP-3-associated osteoarthritis.

Uncovering Novel Extracellular Matrix Transcriptome Alterations in Lesions of Multiple Sclerosis.

The extracellular matrix (ECM) of the central nervous system (CNS) is an interconnected network of proteins and sugars with critical roles in both homeostasis and disease. In neurological diseases, excessive ECM deposition and remodeling impact both injury and repair. CNS lesions of multiple sclerosis (MS), a chronic inflammatory and degenerative disease, cause prominent alterations of the ECM. However, there are a lack of data investigating how the multitude of ECM members change in relation to each other and how this affects the MS disease course. Here, we evaluated ECM changes in MS lesions compared to a control brain using databases generated in-house through spatial mRNA-sequencing and through a public resource of single-nucleus RNA sequencing previously published by Absinta and colleagues. These results underline the importance of publicly available datasets to find new targets of interest, such as the ECM. Both spatial and public datasets demonstrated widespread changes in ECM molecules and their interacting proteins, including alterations to proteoglycans and glycoproteins within MS lesions. Some of the altered ECM members have been described in MS, but other highly upregulated members, including the SPARC family of proteins, have not previously been highlighted. SPARC family members are upregulated in other conditions by reactive astrocytes and may influence immune cell activation and MS disease course. The profound changes to the ECM in MS lesions deserve more scrutiny as they impact neuroinflammation, injury, and repair.

CAFs-Associated Genes (CAFGs) in Pancreatic Ductal Adenocarcinoma (PDAC) and Novel Therapeutic Strategy.

Pancreatic ductal adenocarcinoma (PDAC) is the most aggressive cancer with striking fibrosis, and its mortality rate is ranked second across human cancers. Cancer-associated fibroblasts (CAFs) play a critical role in PDAC progression, and we reviewed the molecular understanding of PDAC CAFs and novel therapeutic potential at present. CAFs-associated genes (CAFGs) were tentatively classified into three categories by stroma specificity representing stroma/epithelia expression ratios (SE ratios). The recent classification using single cell transcriptome technology clarified that CAFs were composed of myofibroblasts (myCAFs), inflammatory CAFs (iCAFs), and other minor ones (e.g., POSTN-CAFs and antigen presenting CAFs, apCAFs). LRRC15 is a myCAFs marker, and myCAFs depletion by diphtheria toxin induces the rapid accumulation of cytotoxic T lymphocytes (CTLs) and therefore augment PDL1 antibody treatments. This finding proposes that myCAFs may be a critical regulator of tumor immunity in terms of PDAC progression. myCAFs are located in CAFs adjacent to tumor cells, while iCAFs marked by PDPN and/or COL14A1 are distant from tumor cells, where hypoxic and acidic environments being located in iCAFs putatively due to poor blood supply is consistent with HIF1A and GPR68 expressions. iCAFs may be shared with SASP (secretion-associated phenotypes) in senescent CAFs. myCAFs are classically characterized by CAFGs induced by TGFB1, while chemoresistant CAFs with SASP may dependent on IL6 expression and accompanied by STAT3 activation. Recently, it was found that the unique metabolism of CAFs can be targeted to prevent PDAC progression, where PDAC cells utilize glucose, whereas CAFs in turn utilize lactate, which may be epigenetically regulated, mediated by its target genes including CXCR4. In summary, CAFs have unique molecular characteristics, which have been rigorously clarified as novel therapeutic targets of PDAC progression.

Validation of the Spanish translation Sheffield Profile for Assessment and Referral for Care (SPARC-Sp) at the Hospital Universitario San Jose of Popayan, Colombia.

OBJECTIVES: We determined the validity and reliability of the Spanish translation Sheffield Profile for Assessment and Referral for Care (SPARC-Sp) questionnaire to identify the palliative care (PC) needs of patients with chronic noncommunicable diseases (NCDs) in Colombia. METHODS: We developed a cross-sectional observational study of scale assessment in adults with the aim of determining the validity and reliability of the SPARC-Sp questionnaire to identify the PC needs of patients with NCDs receiving outpatient or inpatient care at the Hospital Universitario San Jose of Popayan - ESE, Colombia, from 2021 to 2022. RESULTS: We applied a questionnaire consisting of demographic, clinical data, and SPARC-Sp to 507 participants. The constructed model explained 75% of the variance with an adequate fit according to the root mean square residual (0.03), the comparative fit index (0.98), and acceptable reliability (McDonald's total omega 0.4-0.9). Opportunities for improvement are the reformulation and inclusion of particular words to improve the representativeness and clarity of the domains of communication and information, religious, and spiritual issues. SIGNIFICANCE OF RESULTS: This research represents the first validation of SPARC in Spanish. SPARC-Sp is an instrument that allows initiating a conversation of the patient's main needs through a systematic assessment of the patients' main needs. Its psychometric validation demonstrated good fit and acceptable reliability.

Immunohistochemical expression of SPARC in odontogenic keratocysts: a comparative study with other odontogenic cysts.

BACKGROUND: Secreted protein acidic and rich in cysteine (SPARC) has been shown to modulate aggressive behavior in several benign and malignant tumors. Little is known about SPARC expression in odontogenic keratocyst (OKC), an odontogenic cyst with an aggressive nature. To the best of our knowledge, only one study has been investigated the expression of this protein in OKCs. This study aimed to characterize SPARC expression in OKCs. Additionally, to determine whether SPARC is associated with aggressive behavior in OKCs, SPARC expression in OKCs was compared with radicular cysts (RCs), dentigerous cysts (DCs) and calcifying odontogenic cysts (COCs). These odontogenic cysts showed no or less aggressive behavior. METHODS: SPARC expression was evaluated in 38 OKCs, 39 RCs, 35 DCs and 14 COCs using immunohistochemistry. The percentages of positive cells and the intensities of immunostaining in the epithelial lining and the cystic wall were evaluated and scored. RESULTS: Generally, OKCs showed similar staining patterns to RCs, DCs and COCs. In the epithelial lining, SPARC was not detected, except for ghost cells in all COCs. In the cystic wall, the majority of positive cells were fibroblasts. Compared between 4 groups of odontogenic cysts, SPARC expression in OKCs was significantly higher than those of RCs (P < 0.001), DCs (P < 0.001) and COCs (P = 0.001). CONCLUSIONS: A significant increase of SPARC expression in OKCs compared with RCs, DCs and COCs suggests that SPARC may play a role in the aggressive behavior of OKCs.

SPARC in cancer-associated fibroblasts is an independent poor prognostic factor in non-metastatic triple-negative breast cancer and exhibits pro-tumor activity.

Triple-negative breast cancer (TNBC) is the most aggressive breast cancer subtype and lacks specific targeted therapeutic agents. The current mechanistic evidence from cell-based studies suggests that the matricellular protein SPARC has a tumor-promoting role in TNBC; however, data on the clinical relevance of SPARC expression/secretion by tumor and stromal cells in TNBC are limited. Here, we analyzed by immunohistochemistry the prognostic value of tumor and stromal cell SPARC expression in 148 patients with non-metastatic TNBC and long follow-up (median: 5.4 years). We also quantified PD-L1 and PD-1 expression. We detected SPARC expression in tumor cells (42.4%), cancer-associated fibroblasts (CAFs; 88.1%), tumor-associated macrophages (77.1%), endothelial cells (75.2%) and tumor-infiltrating lymphocytes (9.8%). Recurrence-free survival was significantly lower in patients with SPARC-expressing CAFs. Multivariate analysis showed that SPARC expression in CAFs was an independent prognostic factor. We also detected tumor and stromal cell SPARC expression in TNBC cytosols, and in patient-derived xenografts and cell lines. Furthermore, we analyzed publicly available single-cell mRNA sequencing data and found that in TNBC, SPARC is expressed by different CAF subpopulations, including myofibroblasts and inflammatory fibroblasts that are involved in tumor-related processes. We then showed that fibroblast-secreted SPARC had a tumor-promoting role by inhibiting TNBC cell adhesion and stimulating their motility and invasiveness. Overall, our study demonstrates that SPARC expression in CAFs is an independent prognostic marker of poor outcome in TNBC. Patients with SPARC-expressing CAFs could be eligible for anti-SPARC targeted therapy.

Tissue-resident CXCR4+ macrophage as a poor prognosis signature promotes pancreatic ductal adenocarcinoma progression.

Macrophage is an essential part of the tumor immune microenvironment of pancreatic ductal adenocarcinoma. In our study, we explored the CXCR4+ macrophages subset on its prognosis value, immune profile and distinct function in pancreatic cancer progression. Specimens from 102 postoperative pancreatic patients were analyzed by flow cytometry or immune-fluorescence, and the prognostic value of CXCR4+ macrophages infiltration was further determined by Cox regression. In silico analysis on TCGA, ICGC database and single-cell sequencing of pancreatic ductal adenocarcinoma further validated our findings. We found that high CXCR4+ macrophages infiltration was associated with poor overall survival (P < .01) and disease-free survival (P < .05) as an independent factor. CXCR4+ macrophages exhibited an M2 protumor phenotype with high expression of CD206. The function of CXCR4+ macrophages was further analyzed in the murine orthotopic PDAC model with its tumor promotion effect and inhibition of CD8+ T cells. Mechanistic and RNA-seq analysis showed that CXCR4+ macrophages participated in extracellular matrix remodeling procedures and especially secreted SPARC through CXCR4/PI3K/Akt pathway promoting tumor proliferation and migration. Our study reveals that CXCR4+ macrophages infiltration is an indicator of poor prognosis of PDAC and targeting these cells was potentially crucial in immunotherapy of PDAC.

Parallel Evolution of Ameloblastic scpp Genes in Bony and Cartilaginous Vertebrates.

In bony vertebrates, skeletal mineralization relies on the secretory calcium-binding phosphoproteins (Scpp) family whose members are acidic extracellular proteins posttranslationally regulated by the Fam20°C kinase. As scpp genes are absent from the elephant shark genome, they are currently thought to be specific to bony fishes (osteichthyans). Here, we report a scpp gene present in elasmobranchs (sharks and rays) that evolved from local tandem duplication of sparc-L 5' exons and show that both genes experienced recent gene conversion in sharks. The elasmobranch scpp is remarkably similar to the osteichthyan scpp members as they share syntenic and gene structure features, code for a conserved signal peptide, tyrosine-rich and aspartate/glutamate-rich regions, and harbor putative Fam20°C phosphorylation sites. In addition, the catshark scpp is coexpressed with sparc-L and fam20°C in tooth and scale ameloblasts, similarly to some osteichthyan scpp genes. Despite these strong similarities, molecular clock and phylogenetic data demonstrate that the elasmobranch scpp gene originated independently from the osteichthyan scpp gene family. Our study reveals convergent events at the sparc-L locus in the two sister clades of jawed vertebrates, leading to parallel diversification of the skeletal biomineralization toolkit. The molecular evolution of sparc-L and its coexpression with fam20°C in catshark ameloblasts provides a unifying genetic basis that suggests that all convergent scpp duplicates inherited similar features from their sparc-L precursor. This conclusion supports a single origin for the hypermineralized outer odontode layer as produced by an ancestral developmental process performed by Sparc-L, implying the homology of the enamel and enameloid tissues in all vertebrates.

Increased Sparc release from subchondral osteoblasts promotes articular chondrocyte degeneration under estrogen withdrawal.

OBJECTIVE: The incidence of osteoarthritis (OA) in menopausal women is significantly higher than in same-aged men. Investigating the role of subchondral osteoblasts in estrogen deficiency-induced OA may help elucidate the pathological mechanism, providing new insights for the diagnosis and treatment of menopausal OA. METHODS: A classical ovariectomy-induced OA (OVX-OA) rat model was utilized to isolate primary articular chondrocytes and subchondral osteoblasts, which were identified and then cocultured in Transwell. The expression of chondrocyte anabolic and catabolic indicators was evaluated. The differentially expressed proteins in the conditioned medium (CM) of osteoblasts were identified by Liquid Chromatograph-Mass Spectrometer (LC-MS/MS). Normal chondrocytes were treated with osteoblast CM, and then RNA sequencing was performed on the treated chondrocytes. KEGG was used to identify significant enrichment of signaling pathways, and Simple Western was used to verify the expression of related proteins in the signaling pathways. RESULTS: Coculture of OVX-OA subchondral osteoblasts with chondrocytes significantly downregulated the expression of the anabolic indicators and upregulated the expression of the catabolic indicators in chondrocytes. 1,601 proteins were identified in both normal and OVX osteoblast culture supernatants. Protein-protein interaction network analysis revealed that Sparc was one of the hub proteins. The AMPK/Foxo3a signaling pathway of chondrocytes was downregulated by OVX-OA osteoblasts CM. AICAR, the AMPK agonist, partially reversed the catabolic effect of OVX-OA osteoblasts on chondrocytes. CONCLUSIONS: Sparc secreted by OVX-OA subchondral osteoblasts can downregulate the AMPK/Foxo3a signaling pathway of chondrocytes, thereby promoting chondrocyte degeneration.

Coordinated reprogramming of renal cancer transcriptome, metabolome and secretome associates with immune tumor infiltration.

BACKGROUND: Clear cell renal cell carcinoma (ccRCC) is the most common subtype of renal cancer. The molecules (proteins, metabolites) secreted by tumors affect their extracellular milieu to support cancer progression. If secreted in amounts detectable in plasma, these molecules can also serve as useful, minimal invasive biomarkers. The knowledge of ccRCC tumor microenvironment is fragmentary. In particular, the links between ccRCC transcriptome and the composition of extracellular milieu are weakly understood. In this study, we hypothesized that ccRCC transcriptome is reprogrammed to support alterations in tumor microenvironment. Therefore, we comprehensively analyzed ccRCC extracellular proteomes and metabolomes as well as transcriptomes of ccRCC cells to find molecules contributing to renal tumor microenvironment. METHODS: Proteomic and metabolomics analysis of conditioned media isolated from normal kidney cells as well as five ccRCC cell lines was performed using mass spectrometry, with the following ELISA validation. Transcriptomic analysis was done using microarray analysis and validated using real-time PCR. Independent transcriptomic and proteomic datasets of ccRCC tumors were used for the analysis of gene and protein expression as well as the level of the immune infiltration. RESULTS: Renal cancer secretome contained 85 proteins detectable in human plasma, consistently altered in all five tested ccRCC cell lines. The top upregulated extracellular proteins included SPARC, STC2, SERPINE1, TGFBI, while downregulated included transferrin and DPP7. The most affected extracellular metabolites were increased 4-hydroxy-proline, succinic acid, cysteine, lactic acid and downregulated glutamine. These changes were associated with altered expression of genes encoding the secreted proteins (SPARC, SERPINE1, STC2, DPP7), membrane transporters (SLC16A4, SLC6A20, ABCA12), and genes involved in protein trafficking and secretion (KIF20A, ANXA3, MIA2, PCSK5, SLC9A3R1, SYTL3, and WNTA7). Analogous expression changes were found in ccRCC tumors. The expression of SPARC predicted the infiltration of ccRCC tumors with endothelial cells. Analysis of the expression of the 85 secretome genes in > 12,000 tumors revealed that SPARC is a PanCancer indicator of cancer-associated fibroblasts' infiltration. CONCLUSIONS: Transcriptomic reprogramming of ccRCC supports the changes in an extracellular milieu which are associated with immune infiltration. The proteins identified in our study represent valuable cancer biomarkers detectable in plasma.

ROS impairs tumor vasculature normalization through an endocytosis effect of caveolae on extracellular SPARC.

BACKGROUND: The accumulation of reactive oxygen species (ROS) in tumor microenvironment (TME) is an important player for tumorigenesis and progression. We aimed to explore the outcomes of ROS on tumor vessels and the potential regulated mechanisms. METHODS: Exogenous H2O2 was adopted to simulate the ROS setting. Immunofluorescence staining and ultrasonography were used to assess the vascular endothelial coverage and perfusions in the tumors inoculated with Lewis lung cancer (LLC) and melanoma (B16F10) cells of C57BL/6 mice, respectively. ELISA and western-blot were used to detect the expression of secreted acidic and cysteine-rich protein (SPARC) and Caveale-1 in human umbilical vein endothelial cells (HUVEC) extra- and intracellularly. Intracellular translocation of SPARC was observed using electron microscopy and immunofluorescence approaches. RESULT: Under the context of oxidative stress, the pericyte recruitment of neovascularization in mouse lung cancer and melanoma tissues would be aberrated, which subsequently led to the disruption of the tumor vascular architecture and perfusion dysfunction. In vitro, HUVEC extracellularly SPARC was down-regulated, whereas intracellularly it was up-regulated. By electron microscopy and immunofluorescence staining, we observed that SPARC might undergo transmembrane transport via caveale-1-mediated endocytosis. Finally, the binding of SPARC to phosphorylated-caveale-1 was also detected in B16F10 tissues. CONCLUSION: In the oxidative stress environment, neovascularization within the tumor occurs structural deterioration and decreased perfusion capacity. One of the main regulatory mechanisms is the migration of extracellular SPARC from the endothelium to intracellular compartments via Caveolin-1 carriers.

TGF-ß1 upregulates secreted protein acidic and rich in cysteine expression in human granulosa-lutein cells: a potential mechanism for the pathogenesis of ovarian hyperstimulation syndrome.

BACKGROUND: Ovarian hyperstimulation syndrome (OHSS) is a serious complication during in vitro fertilization (IVF) treatment. The upregulation of ovarian transforming growth factor-beta 1 (TGF-ß1) is involved in the development of OHSS. The secreted protein acidic and rich in cysteine (SPARC) is a secreted multifunctional matricellular glycoprotein. Although the regulatory effects of TGF-ß1 on SPARC expression have been reported, whether TGF-ß1 regulates SPARC expression in the human ovary remains unknown. In addition, the role of SPARC in the pathogenesis of OHSS is unclear. METHODS: A steroidogenic human ovarian granulosa-like tumor cell line, KGN, and primary culture of human granulosa-lutein (hGL) cells obtained from patients undergoing IVF treatment were used as experimental models. OHSS was induced in rats, and ovaries were collected. Follicular fluid samples were collected from 39 OHSS and 35 non-OHSS patients during oocyte retrieval. The underlying molecular mechanisms mediating the effect of TGF-ß1 on SPARC expression were explored by a series of in vitro experiments. RESULTS: TGF-ß1 upregulated SPARC expression in both KGN and hGL cells. The stimulatory effect of TGF-ß1 on SPARC expression was mediated by SMAD3 but not SMAD2. The transcription factors, Snail and Slug, were induced in response to the TGF-ß1 treatment. However, only Slug was required for the TGF-ß1-induced SPARC expression. Conversely, we found that the knockdown of SPARC decreased Slug expression. Our results also revealed that SPARC was upregulated in the OHSS rat ovaries and in the follicular fluid of OHSS patients. Knockdown of SPARC attenuated the TGF-ß1-stimulated expression of vascular endothelial growth factor (VEGF) and aromatase, two markers of OHSS. Moreover, the knockdown of SPARC reduced TGF-ß1 signaling by downregulating SMAD4 expression. CONCLUSIONS: By illustrating the potential physiological and pathological roles of TGF-ß1 in the regulation of SPARC in hGL cells, our results may serve to improve current strategies used to treat clinical infertility and OHSS. Video Abstract.