Secretory leukocyte protease inhibitor (SLPI) in cancer pathophysiology: Mechanisms of action and clinical implications.

Cancer is a multifaceted disorder frequently linked to the dysregulation of several biological processes. The SLPI is a multifunctional protein involved in the modulation of immunological response and the inhibition of protease activities. SLPI acts as an inhibitor of proteases, exerts antibacterial properties, and suppresses the transcription of proinflammatory genes through the nuclear factor-kappa B (NF-κB) pathway. The role of this protein as a regulatory agent has been implicated in various types of cancer. Recent research has revealed that SLPI upregulation in cancer cells enhances the metastatic capacity of epithelial malignancies, indicating the deleterious effects of this protein. Furthermore, SLPI interacts intricately with other cancer-promoting factors, including matrix metalloproteinase-2 (MMP-2), MMP-9, the NF-κB and Akt pathways, and the p53-upregulated modulator of apoptosis (PUMA). This review provides an overview of the role of SLPI in cancer pathophysiology, emphasizing its expression in cancer cells and tissues, its potential as a prognostic biomarker, and its therapeutic promise as a target in cancer treatment. The mechanisms of SLPI action in cancer, including its anti-inflammatory effects, regulation of cell proliferation and angiogenesis, and modulation of the tumor microenvironment, have been investigated. The clinical implications of SLPI in cancer have been discussed, including its potential as a diagnostic and prognostic biomarker, its role in chemoresistance, and its therapeutic potential in several types of cancer, such as hepatocellular carcinoma (HCC), colorectal cancer (CRC), pancreatic cancer, head and neck squamous cell carcinoma (HNSCC), ovarian cancer (OvCa), prostate cancer (PC), gastric cancer (GC), breast cancer, and other cancers. In addition, we emphasized the significance of SLPI in cancer, which offers fresh perspectives on potential targets for cancer therapy.

Serum concentrations of secretory leukocyte protease inhibitor and IL6 can predict the onset of sepsis in pyometra bitches.

As onset of sepsis adversely affects the prognosis of canine pyometra, finding biomarkers that would distinguish sepsis status would be useful in the clinical management. Accordingly, we hypothesized that differential expression of endometrial transcripts and circulating concentration of certain inflammatory mediators would discriminate pyometra-led sepsis (P-sepsis+) from those of pyometra without sepsis (P-sepsis-). Bitches with pyometra (n = 52) were classified into P-sepsis+ (n = 28) and P-sepsis- (n = 24) based on vital clinical score and total leukocyte count. A group of non-pyometra bitches (n = 12) served as control. The relative fold changes in the transcripts of IL6, IL8, TNFα, IL10, PTGS2, mPGES1 and PGFS, SLPI, S100A8, S100A12 and eNOS were determined by quantitative polymerase chain reaction. Furthermore, the serum concentrations of IL6, IL8, IL10, SLPI and prostaglandin F2α metabolite (PGFM) were assayed by ELISA. The relative fold changes in S100A12 and SLPI and mean concentrations of IL6 and SLPI were significantly (p < .05) higher in P-sepsis+ than that of P-sepsis- group. Receiver operating characteristic analysis revealed that serum IL6 had a diagnostic sensitivity of 78.6% and a positive likelihood ratio (LR+) of 2.09, at a cut-off value of 15.7 pg/mL to diagnose P-sepsis+ cases. Similarly, serum SLPI had a sensitivity of 84.6% and an LR+ of 2.23, at a cut-off value of 2.0 pg/mL. It was concluded that SLPI and IL6 would serve as putative biomarkers for pyometra-led sepsis in bitches. Monitoring SLPI and IL6 would be a useful adjunct to the established haemato-biochemical parameters in customizing the treatment strategies and arriving at the decision for management of pyometra bitches with critical illness.

Differential expression of inflammatory cytokines, prostaglandin synthases and secretory leukocyte protease inhibitor in the endometrium and circulation in different graded CEH-pyometra in bitch.

Cystic endometrial hyperplasia (CEH)-pyometra (CEH-P) is one of the most common reproductive disorders in bitches, posing a risk to both future fertility and life. The aims of the current study were to elucidate the differential expression patterns of inflammatory mediators at transcript and protein levels in the endometrium and to assess the concentrations of key inflammatory mediators in the peripheral circulation of bitches with different graded CEH-P. A total of 25 client-owned intact mixed breed bitches of 3-10 years presented to the outpatient department of RVP-TVCC of the institute were considered for the study. Of which, 22 cases suggestive of pyometra and 3 cases of CEH obtained during routine elective ovariohysterectomy were subjected to histopathological examination. Uteri were categorized into CEH (n = 3), moderate CEH-P (mCEH-P, n = 9), severe CEH-P (sCEH-P, n = 6) and atrophic pyometra (AT-P, n = 7). A group of age matched (n = 12) bitches without pyometra served as control. Endometrial transcripts such as IL6, IL8, PTGS2, PGFS, and SLPI were expressed differentially in the CEH and CEH-P bitch. In addition, a strong immunoreactivity (IR) of IL6, IL8, PTGS2, and mPGES1 was recorded in the sCEH-P uterus, while expression of IL10 was noticed in AT-P. In circulation, serum IL6 was the most relevant marker with high sensitivity of 96.2% and specificity of 84.6% at a cut off concentration 8.5 pg/mL followed by SLPI with 95.2% sensitivity, and 84.6% specificity at cut off concentration of 1.3 ng/mL. Serum IL10, PGFM and SLPI concentration in the peripheral circulation were 1.5-2.23 fold higher in mCEH-P, 0.87-2.5 fold higher in sCEH-P and 2.9-3.5 fold higher in AT-P than that of control. It is concluded that monitoring the serum concentration of IL6, IL10 and SLPI would be useful adjunct to the established hematobiochemical parameters in the management of pyometra in the bitch with critical illness.

FAPhigh α-SMAlow cancer-associated fibroblast-derived SLPI protein encapsulated in extracellular vesicles promotes ovarian cancer development via activation of PI3K/AKT and downstream signaling pathways.

Ovarian cancer is the most lethal gynecological malignancy worldwide with high metastasis and poor prognosis rates. Cancer-associated fibroblasts (CAFs), a heterogeneous population of cells that constitutes a major component of the tumor microenvironment, secrete extracellular vesicles (EVs) loading with proteins, lipids, and RNAs to promote tumorigenesis. However, the specific roles of CAF-derived proteins contained in EVs in ovarian cancer remain poorly understood at present. Using the gene expression microarray analysis, we identified a list of dysregulated genes between the α-SMA+ CAF and FAP+ CAF subpopulations, from which secretory leukocyte protease inhibitor (SLPI) was chosen for further validation. Quantitative PCR, western blot, immunohistochemistry, and enzyme-linked immunosorbent assays were used to assess SLPI expression in ovarian cancer cells, tissues, CAFs, and EVs. Additionally, we evaluated the effects of exogenous SLPI on proliferation, migration, invasion, and adhesion of ovarian cancer cells in vitro. Our results showed SLPI protein was upregulated in CAFs, particularly in the FAPhigh α-SMAlow CAF subpopulation, and associated with increased tumor grade and decreased overall survival (OS). Importantly, CAF-derived SLPI protein could be encapsulated in EVs for delivery to ovarian cancer cells, thus facilitating cell proliferation, migration, invasion, and adhesion via activating the PI3K/AKT and downstream signaling pathways. Moreover, high plasma expression of SLPI encapsulated in EVs was closely correlated with tumor stage in ovarian cancer patients. Our collective results highlight an oncogenic role of plasma EV-encapsulated SLPI secreted by CAFs in tumor progression for the first time, supporting its potential utility as a prognostic biomarker of ovarian cancer.

High expression of secretory leukocyte protease inhibitor (SLPI) in stage III micro-satellite stable colorectal cancer is associated with reduced disease recurrence.

Secretory leukocyte protease inhibitor (SLPI) is a pleiotropic protein produced by healthy intestinal epithelial cells. SLPI regulates NF-κB activation, inhibits neutrophil proteases and has broad antimicrobial activity. Recently, increased SLPI expression was found in various types of carcinomas and was suggested to increase their metastatic potential. Indeed, we demonstrated that SLPI protein expression in colorectal cancer (CRC) liver metastases and matched primary tumors is associated with worse outcome, suggesting that SLPI promotes metastasis in human CRC. However, whether SLPI plays a role in CRC before distant metastases have formed is unclear. Therefore, we examined whether SLPI expression is associated with prognosis in CRC patients with localized disease. Using a cohort of 226 stage II and 160 stage III CRC patients we demonstrate that high SLPI protein expression is associated with reduced disease recurrence in patients with stage III micro-satellite stable tumors treated with adjuvant chemotherapy, independently of established clinical risk factors (hazard rate ratio 0.54, P-value 0.03). SLPI protein expression was not associated with disease-free survival in stage II CRC patients. Our data suggest that the role of SLPI in CRC may be different depending on the stage of disease. In stage III CRC, SLPI expression may be unfavorable for tumors, whereas SLPI expression may be beneficial for tumors once distant metastases have established.

Levels of secretory leukocyte protease inhibitor expression in acute wounds.

OBJECTIVE: Even with our best practices, we are frequently unable to prevent slow and stalled wound healing-particularly in people with impaired circulation and conditions such as diabetes. As a result, greater insight into the nature of wound healing and alternative treatment approaches is needed. An avenue that may be of particular promise is increasing understanding of the role of secretory leukocyte protease inhibitor (SLPI) as there is evidence that it enhances wound healing, its expression increases in response to inflammation and infection, and it exhibits anti-protease, anti-inflammatory, antiviral antibacterial and antifungal activities. METHOD: The response of SLPI levels to wounding and skin injury was assessed by taking punch skin biopsies from healthy volunteers and assessing the levels of SLPI at the site of injury at the time of wounding (baseline) as well as one, two, three, four, seven, nine and 12 weeks later. RESULTS: A total of 35 volunteers took part in the study. Significant elevations were found: levels of SLPI were greatly increased, 12 times that at baseline, and remained elevated at three weeks despite re-epithelialisation having occurred. CONCLUSION: These findings not only suggest that levels of SLPI rise rapidly following wounding, but that these elevations are sustained, and continue to increase even when re-epithelialisation has occurred. These results suggest that the role and potential benefits of this protease inhibitor deserve further exploration.

Modulation of HIV Replication in Monocyte-Derived Macrophages (MDM) by Host Antiviral Factors Secretory Leukocyte Protease Inhibitor and Serpin Family C Member 1 Induced by Steroid Hormones.

The impact of steroid hormones estrogen and progesterone on human immunodeficiency virus type 1 (HIV-1) replication is well documented. However, the exact mechanism involved in the regulation of HIV-1 replication by estrogen and progesterone is still unclear. In the present study, we wanted to elucidate the molecular mechanisms underlying the modulation of HIV-1 replication by estrogen and progesterone. To achieve this goal, we used real-time quantitative PCR arrays (PCR arrays) to identify differentially expressed host genes in response to hormone treatments that are involved in antiviral responses. Our in vitro results suggest that treatment with high doses of estrogen and progesterone promotes the expression of host antiviral factors Secretory leukocyte protease inhibitor (SLPI) and Serpin family C member 1 (SERPIN C1) among others produced in response to HIV-1 infection. SLPI is an enzyme that inhibits human leukocyte elastase, human cathepsin G, human trypsin, neutrophil elastase, and mast cell chymase. SERPIN C1 is a plasma protease inhibitor that regulates the blood coagulation cascade by the inhibition of thrombin and other activated serine proteases of the coagulation system. A dose dependent downmodulation of HIV-1 replication was observed in monocyte-derived macrophages (MDMs) pre-treated with the two proteins SLPI and SERPIN C1. Further investigations suggests that the host antiviral factors, SLPI and SERPIN C1 act at the pre-integration stage, inhibiting HIV-1 viral entry and leading to the observed downmodulation of HIV-1 replication. Our studies would help identify molecular mechanisms and pathways involved in HIV-1 pathogenesis.

The lower airways microbiome and antimicrobial peptides in idiopathic pulmonary fibrosis differ from chronic obstructive pulmonary disease.

BACKGROUND: The lower airways microbiome and host immune response in chronic pulmonary diseases are incompletely understood. We aimed to investigate possible microbiome characteristics and key antimicrobial peptides and proteins in idiopathic pulmonary fibrosis (IPF) and chronic obstructive pulmonary disease (COPD). METHODS: 12 IPF patients, 12 COPD patients and 12 healthy controls were sampled with oral wash (OW), protected bronchoalveolar lavage (PBAL) and right lung protected sterile brushings (rPSB). The antimicrobial peptides and proteins (AMPs), secretory leucocyte protease inhibitor (SLPI) and human beta defensins 1 and 2 (hBD-1 & hBD-2), were measured in PBAL by enzyme linked immunosorbent assay (ELISA). The V3V4 region of the bacterial 16S rDNA gene was sequenced. Bioinformatic analyses were performed with QIIME 2. RESULTS: hBD-1 levels in PBAL for IPF were lower compared with COPD. The predominant phyla in IPF were Firmicutes, Bacteroides and Actinobacteria; Proteobacteria were among top three in COPD. Differential abundance analysis at genus level showed significant differences between study groups for less abundant, mostly oropharyngeal, microbes. Alpha diversity was lower in IPF in PBAL compared to COPD (p = 0.03) and controls (p = 0.01), as well as in rPSB compared to COPD (p = 0.02) and controls (p = 0.04). Phylogenetic beta diversity showed significantly more similarity for IPF compared with COPD and controls. There were no significant correlations between alpha diversity and AMPs. CONCLUSIONS: IPF differed in microbial diversity from COPD and controls, accompanied by differences in antimicrobial peptides. Beta diversity similarity between OW and PBAL in IPF may indicate that microaspiration contributes to changes in its microbiome.

SLPI suppresses hepatocellular carcinoma progression via endoplasmic reticulum stress induced apoptosis.

Hepatocellular carcinoma (HCC) is one of the most common malignancies worldwide. Secretory leukocyte protease inhibitor (SLPI) has been reported to function as a regulatory factor in several cancers. However, its biological functions and underlying mechanisms in HCC remain to be uncovered. Here, we aimed to explore the effect of SLPI in HCC. In our study, we found that the mRNA and protein expression levels of SLPI were significantly down-regulated in HCC tissues and hepatoma cell lines and low level of SLPI predicted worse survival in our HCC cohorts. In term of function, silencing of SLPI markedly promoted whereas overexpression SLPI suppressed proliferation, migration and invasion capabilities of HCC cells in vitro, and ectopic expression of SLPI inhibited the tumorigenicity of HCC cells in vivo. Mechanistic studies demonstrated that SLPI played a protective role in HCC progression via activating endoplasmic reticulum stress (ER stress)-mediated apoptosis of hepatoma cells, which could be regulated by MAPK signaling pathways. In summary, our findings highlight that SLPI could serve as a potential prognostic biomarker and putative tumor suppressor by enhancing ER stress-induced apoptosis in HCC cells mediated by MAPK signaling pathways, which provides new insights into promising therapeutic targets for HCC treatment.

Non-genetic determinants of malignant clonal fitness at single-cell resolution.

All cancers emerge after a period of clonal selection and subsequent clonal expansion. Although the evolutionary principles imparted by genetic intratumour heterogeneity are becoming increasingly clear1, little is known about the non-genetic mechanisms that contribute to intratumour heterogeneity and malignant clonal fitness2. Here, using single-cell profiling and lineage tracing (SPLINTR)-an expressed barcoding strategy-we trace isogenic clones in three clinically relevant mouse models of acute myeloid leukaemia. We find that malignant clonal dominance is a cell-intrinsic and heritable property that is facilitated by the repression of antigen presentation and increased expression of the secretory leukocyte peptidase inhibitor gene (Slpi), which we genetically validate as a regulator of acute myeloid leukaemia. Increased transcriptional heterogeneity is a feature that enables clonal fitness in diverse tissues and immune microenvironments and in the context of clonal competition between genetically distinct clones. Similar to haematopoietic stem cells3, leukaemia stem cells (LSCs) display heritable clone-intrinsic properties of high, and low clonal output that contribute to the overall tumour mass. We demonstrate that LSC clonal output dictates sensitivity to chemotherapy and, although high- and low-output clones adapt differently to therapeutic pressure, they coordinately emerge from minimal residual disease with increased expression of the LSC program. Together, these data provide fundamental insights into the non-genetic transcriptional processes that underpin malignant clonal fitness and may inform future therapeutic strategies.

Reduced ELANE and SLPI expression compromises dental pulp cell activity.

BACKGROUND: Patients with ELANE variants and severe congenital neutropenia (SCN) commonly develop oral complications. Whether they are caused only by low neutrophil count or the combination of neutropenia and aberrant dental cells is unknown. METHODS: Genetic variant was identified with exome sequencing. Dental pulp cells isolated from the SCN patient with an ELANE mutation were investigated for gene expression, enzyme activity, proliferation, colony formation, wound healing, apoptosis, ROS, attachment, spreading and response to lipopolysaccharide. RESULTS: ELANE cells had diminished expression of ELANE and SLPI and reduced neutrophil elastase activity. Moreover, ELANE cells exhibited impaired proliferation, colony forming, migration, attachment and spreading; and significantly increased ROS formation and apoptosis, corresponding with increased Cyclin D1 and MMP2 levels. The intrinsic levels of TGF-ß1 and TNF-α were significantly increased; however, IL-6, IL-8 and NF-kB1 were significantly decreased in ELANE cells compared with those in controls. After exposure to lipopolysaccharide, ELANE cells grew larger, progressed to more advanced cell spreading stages and showed significantly increased SLPI, TNF-α and NF-kB1 and tremendously increased IL-6 and IL-8 expression, compared with controls. CONCLUSION: This study, for the first time, suggests that in addition to neutropenia, the aberrant levels and functions of ELANE, SLPI and their downstream molecules in pulp cells play an important role in oral complications in SCN patients. In addition, pulp cells with diminished neutrophil elastase and SLPI are highly responsive to inflammation.

Oral secretory leukocyte protease inhibitor (SLPI): Associations with oropharyngeal cancer and treatment outcome.

BACKGROUND: Rates of oropharyngeal cancer (OPC) associated with alcohol & tobacco use have decreased, while human papillomavirus (HPV) associated OPC has increased among men in the US. Secretory leukocyte protease inhibitor (SLPI), detectable in a variety of secretions, has been implicated in cancers of the head and neck, associated with tumor progression and anti-viral activity. Using the recently verified oral gargle specimen, this study aimed to assess the association of salivary SLPI expression with risk of OPC and response to treatment. METHODS: A case-control study design compared levels of salivary SLPI among OPC cases to age and tobacco smoking matched healthy controls. Oral HPV DNA and SLPI was quantified from oral gargle specimens. Logistic regression estimated odds ratios (OR) and 95% confidence intervals (CI) for associations of oral SLPI and risk of OPC and treatment outcomes. RESULTS: In crude and adjusted analyses of 96 OPC cases and 97 age- and smoking-matched controls, OPC was not significantly associated with oral gargle SLPI levels. Among cases, oral SLPI was associated with tonsillectomy (p = 0.018) and among controls oral SLPI was associated with HPV in the oral gargle (p = 0.008). Higher concentrations of SLPI was significantly associated with increased odds of incomplete treatment response (T2: OR: 12.39; 95% CI: 1.44-106.72; T3: OR: 9.86; 95% CI: 1.13-85.90) among all cases, but not among P16+ cases. CONCLUSIONS: Salivary SLPI was not associated with OPC risk but was associated with higher odds of an incomplete treatment response.

Significance of secretory leukocyte peptidase inhibitor in pleural fluid for the diagnosis of benign asbestos pleural effusion.

Secretory leukocyte peptidase inhibitor (SLPI) is a biomarker present in the respiratory tract that protects against tissue destruction and aids in wound healing. We examined whether SLPI in pleural effusion can be used to distinguish benign asbestos pleural effusion (BAPE) from early-stage malignant pleural mesothelioma (MPM) and other diseases. We measured the levels of SLPI, hyaluronic acid (HA), soluble mesothelin-related peptides (SMRP), CCL2, galectin-3, and CYFRA21-1 in 51 patients with BAPE, 37 patients with early-stage MPM, 77 patients with pleural effusions due to non-small-cell lung cancer (LCa), and 74 patients with other pleural effusions. SLPI levels in the pleural fluid of patients with BAPE were significantly lower than those in patients with MPM, LCa, and other pleural effusions (p < 0.0001). The area under the curve (AUC) for SLPI's ability to distinguish BAPE from MPM was 0.902, with a sensitivity of 82.4% and a specificity of 86.5%. This AUC was not only favourable but was better than the AUC for the ability of CYFRA21-1 to distinguish BAPE (0.853). The combination of SLPI and CYFRA21-1 achieved an AUC of 0.965 for the differentiation between BAPE and MPM. Pleural fluid SLPI as well as CYFRA21-1 and HA is useful as a biomarker to diagnose BAPE, which needs to be distinguished from early-stage MPM.

Neutrophil elastase selectively kills cancer cells and attenuates tumorigenesis.

Cancer cell genetic variability and similarity to host cells have stymied development of broad anti-cancer therapeutics. Our innate immune system evolved to clear genetically diverse pathogens and limit host toxicity; however, whether/how innate immunity can produce similar effects in cancer is unknown. Here, we show that human, but not murine, neutrophils release catalytically active neutrophil elastase (ELANE) to kill many cancer cell types while sparing non-cancer cells. ELANE proteolytically liberates the CD95 death domain, which interacts with histone H1 isoforms to selectively eradicate cancer cells. ELANE attenuates primary tumor growth and produces a CD8+T cell-mediated abscopal effect to attack distant metastases. Porcine pancreatic elastase (ELANE homolog) resists tumor-derived protease inhibitors and exhibits markedly improved therapeutic efficacy. Altogether, our studies suggest that ELANE kills genetically diverse cancer cells with minimal toxicity to non-cancer cells, raising the possibility of developing it as a broad anti-cancer therapy.

Reduced Expression of Antimicrobial Protein Secretory Leukoprotease Inhibitor and Clusterin in Chronic Rhinosinusitis with Nasal Polyps.

INTRODUCTION: Antimicrobial peptides and proteins (AMPs) constitute the first line of defense against pathogenic microorganisms in the airway. The association between AMPs and chronic rhinosinusitis with nasal polyps (CRSwNP) requires further investigations. This study is aimed at investigating the expression and regulation of major dysregulated AMPs in the nasal mucosa of CRSwNP. METHODS: The expression of AMPs was analyzed in nasal tissue from patients with eosinophilic (E) CRSwNP and nonECRSwNP and healthy subjects using RNA sequencing. The 10 most abundant AMPs expressed differentially in CRSwNP patients were verified by real-time PCR, and of these, the expression and regulation of secretory leukoprotease inhibitor (SLPI) and clusterin (CLU) were investigated further. RESULTS: The 10 most abundant AMPs expressed differentially in CRSwNP compared to healthy control, regardless of subtypes, included BPIFA1, BPIFB1, BPIFB2, CLU, LTF, LYZ, and SLPI, which were downregulated, and S100A8, S100A9, and HIST1H2BC, which were upregulated. ELISA and immunofluorescence confirmed the decreased expression of SLPI and CLU levels in CRSwNP. SLPI is expressed in both nasal epithelial cells and glandular cells, whereas CLU is mainly expressed in glandular cells. AB/PAS staining further demonstrated that both SLPI and CLU were mainly produced by mucous cells in submucosal glands. Furthermore, the numbers of submucosal glands were significantly decreased in nasal polyp tissue of CRSwNP compared to nasal tissue of controls. SLPI was downregulated by TGF-ß1 and IL-4 in cultured nasal tissues in vitro, while CLU expression was inhibited by TGF-ß1. Glucocorticoid treatment for 2 weeks significantly increased the expression of all downregulated AMPs, but not LYZ. Additionally, budesonide significantly increased the expression of SLPI and CLU in cultured nasal tissues. CONCLUSION: The expression of major antimicrobial proteins is significantly decreased in nasal tissue of CRSwNP. The expression of SLPI and CLU is correlated with the numbers of submucosal glands and regulated by inflammatory cytokines and glucocorticoids.

Overexpression of secretory leukocyte peptidase inhibitor (SLPI) does not modulate experimental osteoarthritis but may be a biomarker for the disease.

OBJECTIVE: Osteoarthritic cartilage destruction can be regulated by the balance between proteases and anti-proteases. Here, we sought to identify novel cellular protease inhibitors associated with osteoarthritis (OA) pathogenesis. METHODS: Candidate molecules were screened from microarray data of chondrocytes treated with OA-associated catabolic factors. The functions of candidate molecules in OA pathogenesis were examined in primary-culture mouse articular chondrocytes and mouse models of OA, such as those stimulated by destabilization of the medial meniscus (DMM) or intra-articular (IA) injection of adenovirus expressing the candidate gene. The value of the selected candidate molecule as a biomarker of OA was examined by measuring its circulating levels in human and mouse blood. RESULTS: Bioinformatic analysis identified secretory leukocyte peptidase inhibitor (SLPI) as a highly upregulated cellular protease inhibitor in chondrocytes treated with pathogenic catabolic factors, including interleukin (IL)-1ß, hypoxia-inducible factor (HIF)-2α, and zinc importer ZIP8. The adenovirus-mediated overexpression of SLPI in joint tissues did not cause any OA-like change or modulate DMM- or HIF-2α-induced experimental OA in mice. SLPI also did not markedly modulate the expression of OA-associated catabolic or anabolic factors in chondrocytes. However, SLPI was specifically upregulated in OA cartilage, and the serum SLPI levels were significantly elevated in human OA patients and experimental OA mice, suggesting that SLPI may be a biomarker of OA. CONCLUSION: Although SLPI is upregulated in OA chondrocytes, it does not appear to per se modulate OA development in mice. However, it may be a potential biomarker of OA in humans and animal models.

Secretory leukocyte protease inhibitor and progranulin as possible regulators of cervical remodeling in pregnancy.

Secretory leukocyte protease inhibitor (SLPI) and progranulin (PGRN) are secretory proteins with an anti-inflammatory property. Their involvement in cervical remodeling in pregnant uterus is not yet elucidated. Thus, this study aimed to explore the significance of SLPI and PGRN in the maintenance of pregnancy by investigating the factors associated with their expression levels at the cervix. Concentrations of SLPI and PGRN proteins were measured in cervical mucus samples collected from asymptomatic pregnant women at 24-26 weeks of gestation (n = 166). The concentrations of those molecules were analyzed with clinical parameters related to risk for preterm delivery (PD). In pregnant mice, we evaluated the effect of lipopolysaccharide-induced inflammation and progesterone effect modulation on cervical mRNA expression of SLPI and PGRN. The cervical PGRN level was significantly lower in women with short cervix (<35 mm) and with a history of threatened PD. In women with short cervix, cervical SLPI concentrations were positively correlated with inflammatory cytokines, interleukin-6 (R2 = 0.75) and interleukin-8 (R2 = 0.71). In pregnant mice, cervical mRNA expressions of PGRN and SLPI were increased in response to progesterone supplementation and were suppressed by a progesterone antagonist, mifepristone. Lipopolysaccharide-induced inflammation caused remarkable upregulation in cervical SLPI mRNA level but not in PGRN. Progesterone and local inflammation are the factors controlling expression levels of PGRN and SLPI at the cervix. The observed relationship of PGRN and SLPI levels in the cervical mucus with PD-related clinical parameters supports that those anti-inflammatory molecules possibly play a significant role in appropriate regulation of cervical remodeling.

SLPI facilitates cell migration by regulating lamellipodia/ruffles and desmosomes, in which Galectin4 plays an important role.

To elucidate the underlying mechanism of secretory leukocyte protease inhibitor (SLPI)-induced cell migration, we compared SLPI-deleted human gingival carcinoma Ca9-22 (ΔSLPI) cells and original (wild-type: wt) Ca9-22 cells using several microscopic imaging methods and gene expression analysis. Our results indicated reduced migration of ΔSLPI cells compared to wtCa9-22 cells. The lamellipodia/dorsal ruffles were smaller and moved slower in ΔSLPI cells compared to wtCa9-22 cells. Furthermore, well-developed intermediate filament bundles were observed at the desmosome junction of ΔSLPI cells. In addition, Galectin4 was strongly expressed in ΔSLPI cells, and its forced expression suppressed migration of wtCa9-22 cells. Taken together, SLPI facilitates cell migration by regulating lamellipodia/ruffles and desmosomes, in which Galectin4 plays an important role.

MiR-16-5p regulates postmenopausal osteoporosis by directly targeting VEGFA.

In this study, we used bioinformatics tools, and experiments with patient tissues and human mesenchymal stem cells (hMSCs) to identify differentially regulated genes (DEGs) and microRNAs (miRNAs) that promote postmenopausal osteoporosis. By analyzing the GSE56815 dataset from the NCBI GEO database, we identified 638 DEGs, including 371 upregulated and 267 downregulated genes, in postmenopausal women with low bone density. Enrichment and protein-protein interaction network analyses showed that TP53, RPS27A, and VEGFA were the top three hub genes with the highest degree of betweenness and closeness centrality. TargetScanHuman and DIANA software analyses and dual luciferase reporter assays confirmed that miR-16a-5p directly targets the 3'UTR of VEGFA. Postmenopausal patients with osteoporosis showed higher miR-16-5p and lower VEGFA levels than those without osteoporosis (n=10 each). VEGFA levels were higher in miR-16-5p knockdown hMSCs and were reduced in miR-16-5p-overexpressing hMSCs. mRNA expression of osteogenic markers, ALP, OCN, and RUNX2, as well as calcium deposition based on Alizarin red staining, correlated inversely with miR-16-5p levels and correlated positively with VEGFA levels. These findings suggest that miR-16-5p suppresses osteogenesis by inhibiting VEGFA expression and is a promising target for postmenopausal osteoporosis therapy.

Genetic activation of Nrf2 reduces cutaneous symptoms in a murine model of Netherton syndrome.

Netherton syndrome is a monogenic autosomal recessive disorder primarily characterized by the detachment of the uppermost layer of the epidermis, the stratum corneum It results from mutations in the SPINK5 gene, which codes for a kallikrein inhibitor. Uncontrolled kallikrein activity leads to premature desquamation, resulting in a severe epidermal barrier defect and subsequent life-threatening systemic infections and chronic cutaneous inflammation. Here, we show that genetic activation of the transcription factor nuclear factor (erythroid-derived 2)-like 2 (Nfe2l2/Nrf2) in keratinocytes of Spink5 knockout mice, a model for Netherton syndrome, significantly alleviates their cutaneous phenotype. Nrf2 activation promoted attachment of the stratum corneum and concomitant epidermal barrier function, and reduced the expression of pro-inflammatory cytokines such as tumor necrosis factor α and thymic stromal lymphopoietin. Mechanistically, we show that Nrf2 activation induces overexpression of secretory leukocyte protease inhibitor (Slpi), a known inhibitor of kallikrein 7 and elastase 2, in mouse and human keratinocytes in vivo and in vitro, respectively. In the Spink5-deficient epidermis, the upregulation of Slpi is likely to promote stabilization of corneodesmosomes, thereby preventing premature desquamation. Our results suggest pharmacological NRF2 activation as a promising treatment modality for Netherton syndrome patients.This article has an associated First Person interview with the first author of the paper.