Inhibiting IL11RA to mitigate hepatic metastasis in skin cutaneous melanoma: Comprehensive insights from in vitro and in vivo investigations.

OBJECTIVE: This study aimed to investigate the role of Interleukin-11 receptor alpha (IL11RA) in skin cutaneous melanoma (SKCM) metastasis to the liver. METHODS: Human SKCM cell lines (A375, A375-MA2, SK-MEL-28, RPMI-7951) and primary dermal fibroblasts (HDFa) were utilized to assess IL11RA expression. IL11RA siRNA was transfected into RPMI-7951 and A375-MA2 cells for Wound healing and Transwell invasion assays. Il11ra knockout (KO) mice and wild-type (WT) mice were injected with B16-F10 cells into the spleen to evaluate hepatic melanoma metastasis. Correlation between IL11RA and MMP family genes was explored using online databases, including LinkedOmics, TIMER (Tumor Immune Estimation Resource), and GEPIA (Gene Expression Profiling Interactive Analysis). RT-qPCR and Western blotting were performed for expression analysis of Mmp2 and Mmp9 in liver tissues of mice. The impact of IL11RA on the STAT3 pathway was investigated in vitro and in vivo. RESULTS: Elevated expression of IL11RA was observed in SKCM cell lines compared to normal cells. IL11RA downregulation significantly inhibited migratory and invasive capabilities of A375-MA2 and RPMI-7951 in vitro. Il11ra gene knockout in mice demonstrated a substantial reduction in hepatic melanoma metastasis. Correlation analyses revealed associations between IL11RA and MMP2/MMP8. Il11ra gene knockout significantly decreased Mmp2 expression while increasing Mmp8 in liver tissues. IL11RA correlated positively with STAT3, and its inhibition led to a suppressed STAT3 pathway in SKCM cells and mouse liver tissue. CONCLUSION: IL11RA plays a crucial role in SKCM metastasis, affecting migratory and invasive abilities. Targeting IL11RA may offer a promising avenue for therapeutic interventions in cutaneous melanoma progression.

Association between Oral Active-Matrix Metalloproteinase-8 Levels and Subretinal Fibrosis among Wet Age-Related Macular Degeneration Patients.

PURPOSE: Periodontitis causes low-grade systemic inflammation and has been associated with elevated active-matrix metalloproteinase (aMMP-8) levels, blood-ocular barrier breakdown and a risk of wet age-related macular degeneration. To assess the association between aMMP-8 levels and macular status among patients with wet age-related macular degeneration (AMD). METHODS: Patients on anti-VEGF treatment for wet AMD were enrolled for oral aMMP-8 rinse test in Mehiläinen Private Hospital, Helsinki, Finland. Macular status was examined from spectral-domain optical coherence tomography (SD-OCT) scans by a medical retina specialist and aMMP-8 levels were analyzed with chairside point-of-care oral rinse (PerioSafe®) test and real-time quantitated by a dentist using the ORALyzer®- reader with a 10 ng/ml cut-off for aMMP-8 activity. RESULTS: Elevated aMMP-8 levels were found in 10 out of 32 patients. Age, gender, anti-VEGF (bevacizumab or aflibercept) distribution, cumulative number of anti-VEGF injections and treatment interval were comparable between patients with aMMP-8 levels below and above the point-of-care level. Macular status differed in regard to aMMP-8 activity; among patients with aMMP-8 levels below the point-of-care subretinal fibrosis was found in 6 out of 22 eyes, whereas among patients with aMMP-8 levels above the point-of-care subretinal fibrosis was found in 8 out of 10 eyes (p = 0.005). Respectively, the mean thickness of subretinal fibrosis at fovea was 19.5 ± 44.1 and 92.3 ± 78.3 µm (p = 0.018). No differences were found in the presence and in the area of geographic atrophy, or fluid distribution, whereas thicknesses of serous pigment epithelial detachment (65.5 ± 99.5 and 12.9 ± 27.9 µm, p = 0.038) and neuroretina (204.2 ± 57.8 µm and 143.0 ± 43.7 µm, p = 0.006) were greater in the eyes of patients with physiological aMMP-8 levels compared to those with elevated aMMP-8 levels. CONCLUSION: Elevated aMMP-8 levels may account for subretinal fibrosis formation in wet AMD.

[Aqueous extract of Epimedium sagittatum mitigates pulmonary fibrosis in mice].

This study aims to investigate the intervention effect of the aqueous extract of Epimedium sagittatum Maxim on the mouse model of bleomycin(BLM)-induced pulmonary fibrosis, so as to provide data support for the clinical treatment of pulmonary fibrosis. Ninety male C57BL/6N mice were randomized into normal(n=10), model(BLM, n=20), pirfenidone(PFD, 270 mg·kg~(-1), n=15), and low-, medium-, and high-dose E. sagittatum extract(1.67 g·kg~(-1), n=15; 3.33 g·kg~(-1), n=15; 6.67 g·kg~(-1), n=15) groups. The model of pulmonary fibrosis was established by intratracheal instillation of BLM(5 mg·kg~(-1)) in the other five groups except the normal group, which was treated with an equal amount of normal saline. On the day following the modeling, each group was treated with the corresponding drug by gavage for 21 days. During this period, the survival rate of the mice was counted. After gavage, the lung index was calculated, and the morphology and collagen deposition of the lung tissue were observed by hematoxylin-eosin(HE) and Masson staining, respectively. The levels of reactive oxygen species(ROS) in lung cell suspensions were measured by flow cytometry. The levels of glutathione peroxidase(GSH-Px), total superoxide dismutase(T-SOD), and malondialdehyde(MDA) the in lung tissue were measured. Terminal-deoxynucleoitidyl transferase-mediated nick-end labeling(TUNEL) was employed to examine the apoptosis of lung tissue cells. The content of interleukin-6(IL-6), chemokine C-C motif ligand 2(CCL-2), matrix metalloproteinase-8(MMP-8), transforming growth factor-beta 1(TGF-ß1), alpha-smooth muscle actin(α-SMA), E-cadherin, collagen Ⅰ, and fibronectin in the lung tissue was measured by enzyme-linked immunosorbent assay(ELISA). The expression levels of F4/80, Ly-6G, TGF-ß1, and collagen Ⅰ in the lung tissue were determined by immunohistochemistry. The mRNA levels of CCL-2, IL-6, and MMP-7 in the lung tissue were determined by qRT-PCR. The content of hydroxyproline(HYP) in the lung tissue was determined by alkaline hydrolysation. The expression of α-SMA and E-cadherin was detected by immunofluorescence, and the protein levels of α-SMA, vimentin, E-cadherin in the lung tissue were determined by Western blot. The results showed the aqueous extract of E. sagittatum increased the survival rate, decreased the lung index, alleviated the pathological injury, collagen deposition, and oxidative stress in the lung tissue, and reduced the apoptotic cells. Furthermore, the aqueous extract of E. sagittatum down-regulated the protein levels of F4/80 and Ly-6G and the mRNA levels of CCL-2, IL-6, and MMP-7 in the lung tissue, reduced the content of IL-6, CCL-2, and MMP-8 in the alveolar lavage fluid. In addition, it lowered the levels of HYP, TGF-ß1, α-SMA, collagen Ⅰ, fibronectin, and vimentin, and elevated the levels of E-cadherin in the lung tissue. The aqueous extract of E. sagittatum can inhibit collagen deposition, alleviate oxidative stress, and reduce inflammatory response by regulating the expression of the molecules associated with epithelial-mesenchymal transition, thus alleviating the symptoms of bleomycin-induced pulmonary fibrosis in mice.

Altered Peripheral Blood Gene Expression in Childhood Cancer Survivors With Anthracycline-Induced Cardiomyopathy - A COG-ALTE03N1 Report.

Background Anthracycline-induced cardiomyopathy is a leading cause of premature death in childhood cancer survivors, presenting a need to understand the underlying pathogenesis. We sought to examine differential blood-based mRNA expression profiles in anthracycline-exposed childhood cancer survivors with and without cardiomyopathy. Methods and Results We designed a matched case-control study (Children's Oncology Group-ALTE03N1) with mRNA sequencing on total RNA from peripheral blood in 40 anthracycline-exposed survivors with cardiomyopathy (cases) and 64 matched survivors without (controls). DESeq2 identified differentially expressed genes. Ingenuity Pathway Analyses (IPA) and Gene Set Enrichment Analyses determined the potential roles of altered genes in biological pathways. Functional validation was performed by gene knockout in human-induced pluripotent stem cell-derived cardiomyocytes using CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9) technology. Median age at primary cancer diagnosis for cases and controls was 8.2 and 9.7 years, respectively. Thirty-six differentially expressed genes with fold change ≥±2 were identified; 35 were upregulated. IPA identified "hepatic fibrosis" and "iron homeostasis" pathways to be significantly modulated by differentially expressed genes, including toxicology functions of myocardial infarction, cardiac damage, and cardiac dilation. Leading edge analysis from Gene Set Enrichment Analyses identified lactate dehydrogenase A (LDHA) and cluster of differentiation 36 (CD36) genes to be significantly upregulated in cases. Interleukin 1 receptor type 1, 2 (IL1R1, IL1R2), and matrix metalloproteinase 8, 9 (MMP8, MMP9) appeared in multiple canonical pathways. LDHA-knockout human-induced pluripotent stem cell-derived cardiomyocytes showed increased sensitivity to doxorubicin. Conclusions We identified differential mRNA expression profiles in peripheral blood of anthracycline-exposed childhood cancer survivors with and without cardiomyopathy. Upregulation of LDHA and CD36 genes suggests metabolic perturbations in a failing heart. Dysregulation of proinflammatory cytokine receptors IL1R1 and IL1R2 and matrix metalloproteinases, MMP8 and MMP9 indicates structural remodeling that accompanies the clinical manifestation of symptomatic cardiotoxicity.

Effect of intracanal cryotherapy on the inflammatory cytokine, proteolytic enzyme levels and post-operative pain in teeth with asymptomatic apical periodontitis: A randomized clinical trial.

AIM: To investigate the changes in the biomarker levels related to inflammation and tissue destruction in the periapical exudate of mandibular pre-molar teeth with asymptomatic apical periodontitis after receiving intracanal cryotherapy, to compare cryotherapy and control groups in terms of analgesic intake, interappointment, and post-operative pain and evaluate the correlation between biomarker levels and interappointment pain. METHODOLOGY: Mandibular pre-molar teeth of 44 patients aged 18-35 years, diagnosed with asymptomatic apical periodontitis, were root canal treated in two visits (registered as NCT04798144). Baseline periapical exudate samples were obtained, and the patients were assigned to either control or intracanal cryotherapy group according to the final irrigation with distilled water either at room temperature or 2.5°C. The canals were dressed with calcium hydroxide. In the second visit, the calcium hydroxide was removed with passive ultrasonic irrigation, and the periapical exudate was sampled again. IL-1ß, IL-2, IL-6, IL-8, TNF-α, PGE2 and MMP-8 levels were determined with ELISA. Post-operative pain levels were recorded for 6 days following both visits using a visual analogue scale. Data were analysed using t-test, the Mann-Whitney U test and correlation tests. RESULTS: There was a significant correlation between the pain scores reported after first visit and IL-1ß and PGE2 levels (p < .05). IL-1ß, IL-2 and IL-6 levels showed no significant difference in the cryotherapy group (p > .05), while they significantly increased in the control group (p < .05). There was a decrease in IL-8, TNF-α, PGE2 and MMP-8 levels, however, the difference was not significant (p > .05). Pain scores were significantly lower in the cryotherapy group for the first 3 days (p < .05), except for 24th hours (p > .05). CONCLUSIONS: The positive correlation between interappointment pain and IL-1ß and PGE2 levels might indicate that these biomarker levels can be used to predict the severity of post-operative pain. Intracanal cryotherapy was effective in reducing post-operative pain in the short term in teeth with asymptomatic apical periodontitis. Cryotherapy prevented an increase in IL-1ß, IL-2 and IL-6 levels compared with the control group.

Therapeutic efficacy of adjunctive photodynamic therapy in the treatment of denture stomatitis.

BACKGROUND: The present report assessed the efficacy of curcumin-mediated photodynamic therapy (CUR-mediated PDT) as an adjunct to antifungal gel treatment by evaluating the salivary interleukin-6 (IL-6) and matrix metalloproteinases-8 (MMP-8) levels together with Candida species counts in denture stomatitis (DS) patients. METHODS: In total, 50 DS subjects were randomly categorized into 2 groups: Group-1: subjects who received the antifungal gel treatment and Group-2: participants who received CUR-mediated PDT. The Sabourad Dextrose Agar and CHROMAgar were utilized for evaluating Candida species counts, while the Enzyme-Linked Immunosorbent Assay was employed to estimate the salivary levels of IL-6 and MMP-8. All clinical evaluations were performed at the baseline, 1 month, and 2 months. RESULTS: In total, group-2 subjects showed a significant decrease in Candida albicans (C. albicans) counts on both follow-ups (i.e., 1-month and 2-month) than group-1 participants. C. krusei count also reduced in group-2 subejcts than group-1 participants at the 2nd follow-up as compared to the baseline, nevertheless, a slight increase in C. krusei count was noticed in group-2 subjects at the 2nd follow-up than the 1st follow-up. The salivary IL-6 and MMP-8 levels in both groups reduced significantly at both follow-ups than the baseline. According to the stepwise logistic regression analysis, no statistically significant correlation was observed between Candida species count and other parameters such as age and gender of the patient, duration of DS, and frequency of treatment(s). CONCLUSION: CUR-mediated PDT is an efficaciousness therapeutic modality for alleviating Candida species counts on the surface of denture and the palatal mucosa, as well as improving the salivary IL-6 and MMP-8 levels in DS patients.

In-silico Validation of the Proposed Treatment Strategy of Periodontitis.

OBJECTIVE: The present study aims to assess a proposed treatment approach or therapy for periodontitis by using the in-silico technique. The proposed treatment strategy offers a singular vehicular system consisting of minocycline (antibiotic), celecoxib (selective COX-II inhibitor), doxycycline hyclate (matrix metalloproteinase inhibitor), and hydroxyapatite (osteogenic agent). MATERIAL AND METHODS: Molecular docking studies of drugs were performed using Maestro version 9.4 software Schrödinger, and 3-Dimensional Crystallographic X-ray protein structures of targeted proteins were downloaded from RCSB protein data bank in .pdb file format. These agents were docked, and their affinities towards the receptors/protein/enzyme were calculated. Furthermore, their affinities were compared with the standard drug. RESULTS: The study suggests that minocycline and metronidazole possess equal affinity towards the RGPB and Inlj protein of P.gingivalis. Celecoxib, a well-known inhibitor of the COX-II enzyme, showed very high affinity. Selective inhibitor of MMP-8 possessed higher affinity than doxycycline, whereas CMT-3 showed equal affinity as doxycycline for MMP-13. Similarly, hydroxyapatite and simvastatin also showed a comparatively similar affinity for osteopontin receptor. CONCLUSION: Based upon molecular docking results, it can be concluded that the proposed treatment strategy would be a suitable approach for periodontitis and all the selected therapeutic agents have potential similar to the standard drugs, thereby constituting a reliable system for periodontitis.

An Injectable Containing Morphine, Ropivacaine, Epinephrine, and Ketorolac Is Not Cytotoxic to Articular Cartilage Explants From Degenerative Knees.

PURPOSE: The purpose of this study was to determine the effects of a multidrug injectate containing morphine, ropivacaine, epinephrine, and ketorolac, commonly referred to as the "Orthococktail," on cartilage tissue viability and metabolic responses using an established in vitro model. METHODS: With institutional review board approval and informed patient consent, tissues normally discarded after total knee arthroplasty (TKA) were recovered. Full-thickness cartilage explants (n = 72, Outerbridge grade 1 to 3) were created and bisected. Paired explant halves were treated with either 1 mL Orthococktail or 1 mL of saline and cultured for 8 hours at 37°C, with 0.5 mL of the treatment being removed and replaced with tissue culture media every hour. Explants were cultured for 6 days, and media were changed and collected on days 3 and 6. After day 6, tissues were processed for cell viability, weighed, and processed for histologic grading. Outcome measures were compared for significant differences between treated and untreated samples. RESULTS: There were no significant differences in cartilage viability between control and Orthococktail-treated samples across a spectrum of cartilage pathologies. Orthococktail treatment consistently resulted in a significant decrease in the release of PGE2, MCP-1, MMP-7, and MMP-8 on day 3 of culture and PGE2, MMP-3, MMP-7, and MMP-8 on day 6 of culture, compared with saline controls. CONCLUSION: The results of the present study indicate that an Orthococktail injection composed of morphine, ropivacaine, epinephrine, and ketorolac is associated with a transient decrease in degradative and inflammatory mediators produced by more severely affected articular cartilage and may mitigate perioperative joint pain such that postoperative narcotic drug use could be reduced. CLINICAL RELEVANCE: The Orthococktail solution used in this study may be a safe intraoperative, intra-articular injection option for patients undergoing joint arthroplasty and other joint preservation surgical procedures.

Evaluation of Wound Closure Rates Using a Human Fibroblast-derived Dermal Substitute Versus a Fetal Bovine Collagen Dressing: A Retrospective Study.

Diabetic foot ulcers (DFUs) are associated with an increased risk for serious and costly outcomes such as osteomyelitis, amputation, and hospitalization. PURPOSE: A retrospective study was conducted to evaluate the proportion of patients healed and time to healing of DFUs treated with a human fibroblast-derived dermal substitute (HFDS) or a fetal bovine collagen dressing (FBCD). METHODS: Data from patients with a DFU who received the first treatment in 2014 were extracted from the electronic record database of 93 wound care centers. Baseline demographics (eg, age, gender, body mass index, and number of wounds); wound location, size, and duration; and wound-specific information such as wound size and number of and interval between applications were obtained. Study criteria stipulated patients who received at least one treatment in 2014 with HFDS or FBCD on a DFU with location coded as foot, toe, heel, metatarsal head, toe web space, toe amputation site, or transmetatarsal amputation site; ulcer size ≥1 cm2 to <20 cm2; and ulcer area reduction ≤50% in the 28 days before the first treatment with HFDS or FBCD were eligible for inclusion. Wounds that received an alternate skin substitute treatment up to 28 days before or concurrent with the first HFDS or FBCD treatment or if patient data that lacked baseline or follow-up wound area measurement were excluded. Deidentified data were extracted directly into data files and transferred to a third-party data management and statistical group for analysis. The frequency of DFUs achieving wound closure (defined as area ≤0.25 cm2) by weeks 12 and 24 and median time to wound closure of wounds that healed were analyzed. Baseline characteristics were compared using 2-sample t tests for continuous variables and 2-tailed Fisher's exact tests for difference in proportions between treatments. Frequency of and median time to wound closure were determined by Cox proportional hazards analysis. The frequency of wounds closed at 12 and 24 weeks, median time to wound closure, hazard ratio with 95% confidence interval, and P value were estimated from the Cox model. Statistical significance was defined as P <.05. RESULTS: Records showed 206 patients with 208 DFUs received treatment (108 HFDS, mean age 60.2 years, mean wound duration 8.8 months; 100 FBCD, mean age 65.2 years, mean wound duration 12.8 months) and were included. Mean number of treatment applications was 4.5 and 2.4 for HFDS and FBCD, respectively. After 12 and 24 weeks 44 (41%) and 69 (64%) of HFDS-treated wounds, respectively, and 21 (21%) and 43 (43%) of FBCD-treated wounds, respectively, were healed (at 12 weeks, P = .03; at 24 weeks, P = .03, log rank 2-tailed test, unadjusted). Median time to wound closure for HFDS and FBCD was 14.6 and 25 weeks, respectively (P = .03; log rank, 2-tailed test; Kaplan-Meier analysis). HFDS treatment significantly increased the probability of wound healing compared to FBCD treatment in the Cox proportional hazards analysis after adjusting for treatment terms, baseline wound area, baseline wound duration, baseline wound depth, wound location, and patient age at first treatment (HR = 1.77; 95% CI: 1.06-2.97; P = .03). CONCLUSION: DFU wounds are more likely to heal when treated with HFDS than with FBCD as used by facilities in this database. Studies examining the efficacy, cost-effectiveness, and patient-centered outcomes of these treatments is warranted. .

Truncated active human matrix metalloproteinase-8 delivered by a chimeric adenovirus-hepatitis B virus vector ameliorates rat liver cirrhosis.

BACKGROUND: Liver cirrhosis is a potentially life-threatening disease caused by progressive displacement of functional hepatocytes by fibrous tissue. The underlying fibrosis is often driven by chronic infection with hepatitis B virus (HBV). Matrix metalloproteinases including MMP-8 are crucial for excess collagen degradation. In a rat model of liver cirrhosis, MMP-8 delivery by an adenovirus (Ad) vector achieved significant amelioration of fibrosis but application of Ad vectors in humans is subject to various issues, including a lack of intrinsic liver specificity. METHODS: HBV is highly liver-specific and its principal suitability as liver-specific gene transfer vector is established. HBV vectors have a limited insertion capacity and are replication-defective. Conversely, in an HBV infected cell vector replication may be rescued in trans by the resident virus, allowing conditional vector amplification and spreading. Capitalizing on a resident pathogen to help in its elimination and/or in treating its pathogenic consequences would provide a novel strategy. However, resident HBV may also reduce susceptibility to HBV vector superinfection. Thus a size-compatible truncated MMP-8 (tMMP8) gene was cloned into an HBV vector which was then used to generate a chimeric Ad-HBV shuttle vector that is not subject to superinfection exclusion. Rats with thioacetamide-induced liver cirrhosis were injected with the chimera to evaluate therapeutic efficacy. RESULTS: Our data demonstrate that infectious HBV vector particles can be obtained via trans-complementation by wild-type virus, and that the tMMP8 HBV vector can efficiently be shuttled by an Ad vector into cirrhotic rat livers. There it exerted a comparable beneficial effect on fibrosis and hepatocyte proliferation markers as a conventional full-length MMP-8Ad vector. CONCLUSIONS: Though the rat cirrhosis model does not allow assessing in vivo HBV vector amplification these results advocate the further development of Ad-HBV vectors for liver-specific gene therapy, including and perhaps particularly for HBV-related disease.