MMP-2 Activity Research Articles

Hsa-circ-0006091 modulates the proliferation of hepatocellular carcinoma via the miR-622/CCNB1 axis.

Hepatocellular carcinoma (HCC) is a common type of cancer. We hypothesize that circular RNA-0006091 (circ-0006091) affects the progression of HCC. The study aims to investigate the effect of circ-0006091 in HCC cells. The levels of circ-0006091, microRNA-622 (miR-622), and cyclin B1 (CCNB1) were assayed using qRT-PCR and western blotting. The metastasis of the HCC cells was measured with wound healing and transwell assays. The protein expression levels of MMP-2 and MMP-9 were assayed with western blotting. Dual-luciferase reporter and RNA-pulldown assays were used to determine the link between miR-622 and circ-0006091 or CCNB1. Mice-based tests were used to determine the effect of circ-0006091 on the proliferation of HCC cells. The levels of circ-0006091 and CCNB1 were increased in the HCC cells, but miR-622 was down-regulated. Deficiency of circ-0006091 reduced the metastasis of the HCC cells, and silencing of circ-0006091 decreased the activities of MMP-2 and MMP-9 in the same cells. Circ-0006091 modulated the CCNB1 level in the HCC cells via miR-622. Silencing of circ-0006091 suppressed the proliferation of the HCC cells in vivo. Circ-0006091 regulated HCC cell metastasis via the miR-622/CCNB1 axis, a possible therapeutic target in managing HCC.

Correlations between different protein species of oral rinse MMP-8 and patient-related factors.

The aim of this study is to examine correlations between different oral rinse matrix metalloproteinase (MMP)-8 protein species in western blot (WB) analysis, quantitative MMP-8 measurements, and patient-related factors. Elevated activated MMP-8 (aMMP-8) associate with periodontitis and a diagnostic point-of-care technology has been developed based on aMMP-8. In WB, different MMP-8 protein species can be analyzed. Relative abundancyof fragmented 20-25 kDa forms in WB has been associated with and reflects MMP-8 activation and related fragmentation and elevated quantitative aMMP-8 measurements. A random sample of 192 participants from a periodontal disease screening study was used for this study. Oral rinse samples for biomarker analyses were collected before clinical periodontal examinations. aMMP-8 immunofluorometric (IFMA) and WB analysis (utilizing the same monoclonal antibody, 8708), polymorphonuclear leukocyte (PMN) elastase activity test and tissue inhibitor of metalloproteinases (TIMP)-1 ELISA levels were performed from the oral rinse samples. Distinct MMP-8 protein species were differentiated in the WB analysis. Principal component (PC) analysis was conducted to explore correlation patterns between the different species. Adjusted correlation analysis between the extracted PCs of WB and aMMP-8 IFMA levels and multilevel regression analysis were conducted to explore if the other periodontal disease-related biomarkers and clinical surrogate measures and patient-related factors are co-variating with the extracted components. Distinct correlation patterns between the MMP-8 protein species were observed. The first four PCs explained 89% of the whole variance in PC analysis. Statistically significant correlation (p < 0.05) were observed as follows: PC1 positively with 21 kDa (r = .69) and 25 kDa fragments (r = .55) and negatively with 150 kDa complexes (r = -.46). PC2 correlated with 45 (r = .70) and 55 kDa (r = .65) activated forms, PC3 with 70-80 kDa latent proforms (r = .63) and 90-100 kDa complexes (r = .67), and PC4 with 35 kDa fragments (r = .81). There were significant correlations between quantitative (IFMA) aMMP-8 measurements and PC1 (p < 0.001), PC2 (<0.05) and PC3 (<0.05) but not with PC4. In multilevel regression models age, PMN elastase activity, TIMP-1 levels, and a number of 4-5 mm periodontal pockets were associated with PC1, nonsmoking with PC2, ageand PMN elastase activity with PC3, and age and smoking with PC4. Relative abundancy of fragmented 21-25 kDa protein species was correlated with the quantitative aMMP-8 (IFMA) measurements, which is in line with previous results. Different patient-related factors (smoking, age, proteolytic activity) may modify the formation of different MMP-8 protein species in oral rinse samples and may cause variability in quantitative aMMP-8 measurement.

Early growth response 2, a novel target of pelvic organ prolapse, is highly expressed in anterior vaginal wall tissues with pelvic organ prolapse.

Pelvic organ prolapse (POP) is a common disorder among women that negatively affects women's quality of life. Early growth response 2 (EGR2) is a transcription factor that regulates cell growth. The present study aimed to explore the role of EGR2 in POP progression and provided a new target for the treatment and prevention of POP. Firstly, we extracted primary vaginal anterior wall fibroblasts from POP tissues and non-POP tissues and then constructed an EGR2-silencing lentivirus for further study. Immunoblotting, qPCR, TUNEL assay, CCK-8 assay, dual luciferase assay, and ELISA assay were carried out. EGR2 expression was much higher in POP tissues than in control tissues, and EGR2 expression positively correlated with cytokine signaling 3 (SOCS3) expression. Knockdown of EGR2 increased cell proliferation, upregulated PCNA expression, and reduced apoptosis in POP fibroblasts. Moreover, we found that the knockdown of EGR2 increased COL1A1, COL3A1, and Elastin expression and decreased MMP2 and MMP9 activities, and knockdown of EGR2 increased TGF-β/Smad pathway activity in POP fibroblasts. Interestingly, the results of dual luciferase assay demonstrated that EGR2 was able to increase SOCS3 transcriptional activity. EGR2 knockdown alleviated the apoptosis of POP fibroblasts by reducing SOCS3 expression and improving the proliferation and collagen synthesis of POP fibroblasts. Overall, our study illustrated that EGR2 was highly expressed in POP tissues, and knockdown of EGR2 alleviated apoptosis and reduced matrix degradation in POP fibroblasts. This study might provide a new insight into the pathogenesis of POP.

The combination of cigarette smoke and solar rays causes effects similar to skin aging in a bilayer skin model

Skin aging is a multifactorial process influenced by internal and external factors. The contribution of different environmental factors has been well established individually in the last few years. On the one hand, man is rarely exposed to a single factor, and on the other hand, there is very little knowledge about how these extrinsic factors may interact with each other or even how the skin may react to chronic exposure. This study aimed to evaluate the effect on skin aging of a chronic co-exposure of tissue-engineered skin substitutes to cigarette smoke extract (CSE) and solar simulator light (SSL). Skin substitutes were reconstructed according to the self-assembly method and then exposed to CSE followed by irradiation with SSL simultaneously transmitting UVA1, visible light and infrared. When skin substitutes were chronically exposed to CSE and SSL, a significant decrease in procollagen I synthesis and the inhibition of Smad2 phosphorylation of the TGF-β signaling pathway were observed. A 6.7-fold increase in MMP-1 activity was also observed when CSE was combined with SSL, resulting in a decrease in collagen III and collagen IV protein expression. The secretory profile resulting from the toxic synergy was investigated and several alterations were observed, notably an increase in the quantities of pro-inflammatory cytokines. The results also revealed the activation of the ERK1/2 (3.4-fold) and JNK (3.3-fold) pathways. Taken together, the results showed that a synergy between the two environmental factors could provoke premature skin aging.

Combretastatin A-4 suppresses the invasive and metastatic behavior of glioma cells and induces apoptosis in them: in-vitro study.

The most common primary brainmalignancy, glioblastoma multiforme, is tremendously resistant to conventionaltreatmentsdue to its potency for metastasis to surrounding brain tissue. Temozolomide is a chemotherapeutic agent that currently is administrated during the treatment procedure. Studies have attempted to investigate new agents with higher effectiveness and fewer side effects. Combretastatin A-4 (CA-4), a natural compound derived from Combretum caffrum, has been recently considered for its potent antitumor activities in a wide variety of preclinical solid tumor models. Our findings have shown that CA-4 exerts potent anti-proliferative and apoptotic effects on glioma cells, and ROS generation may be involved in these cellular events. CA-4 has imposed G2 arrest in U-87 cells. We also observed that CA-4 significantly reduced the migration and invasion capability of U-87 cells. Furthermore, the gene expression and enzyme activity of MMP-2 and MMP-9 were significantly inhibited in the presence of CA-4. We also observed a considerable decrease in PI3K and Akt protein expression following treatment with CA-4. In conclusion, our findings showed significant apoptogenic and anti-metastatic effects of CA-4 on glioma cells and also suggested that the PI3K/Akt/MMP-2/-9 and also ROS pathway might play roles in these cellular events.

Effect of etchant containing an Urushiol monomer from lacquer sap on dentin biostability and bonding performance.

Objectives: This study aimed to evaluate the effectiveness of urushiol as an additive to surface acid etchant on dentin structure, by assessing the biostability of dentin, and determine the bonding strengths of dentin and enamel to the composite in the complicated oral microecology. Methods: Etchants with different concentrations of urushiol (0.5, 1, or 3 wt%) were formulated and tested for their bonding performance. Demineralized dentin beams that were etched with experimental etchants were incubated in simulated body fluid solutions by evaluating the weight decrement after 1month. The effects of urushiol on dentin and matrix metalloproteinases were confirmed by scanning electron microscopy (SEM). Moreover, the antibiotic actions of urushiol on the common cariogenic bacteria Streptococcus mutans, Streptococcus sanguinis, and Streptococcus gordonii as well as the biofilm were evaluated, and its effect on bacterial morphology was observed by scanning electron microscopy. Finally, enamel and dentin specimens were prepared from human molars to determine the depth of demineralization by the etchants and the relationship with the resin bond strengths to enamel and dentin (μTBS) and the morphology of the bonding interface. Results: Urushiol could interact with dentine and inhibit collagenase activity, resulting in biostable dentine. The application of the etchants containing 0.5, 1, or 3 wt% urushiol significantly improved the durability of the dentin bonding interface with its instinctive antibacterial property (p < 0.05). Conclusion: Urushiol not only improves dentin stability by interacting with collagen and inactivating MMP activity but also plays a role in the antibacterial effects in the complicated oral microecology. The effectiveness of urushiol etchant prolongs the longevity of bonded dental restorations without compromising clinical operation time.

Exosomes isolated from metabolically unhealthy normal weight and overweight phenotypes deteriorate the ER/PR positive breast cancer behavior.

Obesity has been linked to a higher risk of postmenopausal breast cancer Yet, research indicates an opposite correlation between obesity and premenopausal breast cancer risk. Various obesity phenotypes based on metabolic health could play a significant part. This study aims to assess how plasma exosomes taken from women with varying obesity phenotypes impact MCF-7 cell migration, matrix metalloproteinase-2 activity, and apoptosis. The characterization of isolated exosomes and their internalization into MCF-7 cells was evaluated. The treatment of MCF-7 cells with exosomes isolated from different groups was done. Migration, the activity of MMP-2, mRNA expression of Bax and Bcl-2, protein expression of p-53 and Thr55 p-p53, and apoptosis were assessed. Isolated exosomes from unhealthy obese individuals increase MCF-7 cell migration. Regarding MMP activities, unhealthy normal weight and overweight and healthy obese groups isolated exosomes increase the MMP-2 activity than the treated group with exosomes isolated from counterpart groups. Furthermore, unhealthy normal weight and overweight and healthy obese obtained exosomes decrease apoptosis compared to counterpart groups. Altogether, plasma exosomes derived from both unhealthy individuals with normal weight and overweight status, as well as those with unhealthy obesity, negatively impacted the behavior of estrogen/progesterone receptor-positive breast cancer cells. The online version contains supplementary material available at 10.1007/s40200-023-01295-1.

Anti-Inflammatory Effects of Boric Acid in Treating Knee Osteoarthritis: Biochemical and Histopathological Evaluation in Rat Model.

This study aimed to examine the anti-inflammatory properties of boric acid (BA) in treating knee osteoarthritis (KOA) in rats, evaluating its biochemical and histopathological therapeutic effects.A KOA rat model was induced by injecting monosodium iodoacetate into the knee joint. Random assignment was performed for the experimental groups as follows: group-1(control), group-2(KOA control), group-3 (BA:4mg/kg, orally), group-4(BA:10mg/kg, orally), group-5(BA:4mg/kg, intra-articularly), and group-6(BA:10mg/kg, intra-articularly). The rats received 100 µL of BA intra-articularly on days 1, 7, 14, and 21 or 1mL orally once a day (5days/week) for 4weeks. Serum levels of interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), and activityofmatrix metalloproteinase-13 (MMP-13) were measured. Histopathological and immunohistochemical analyses were performed on knee joint samples using specific antibodies for IL-1β, TNF-α, MMP-13, and nitric oxide synthase-2 (NOS-2).Group-2 exhibited higher serum IL-1β and TNF-α levels and MMP-13 activity than group-1 (P < 0.05). However, IL-1β and TNF-α levels and MMP-13 activity were lower in all treatment groups than in group-2, with statistically significant reductions observed in groups-4, 5, and 6. Histopathologically, group-2 displayed joint space narrowing, cartilage degeneration, and deep fissures. Groups-5 and 6 demonstrated significant joint space enlargement, articular cartilage tissue regeneration, and immunostaining patterns similar to those in group-1. Immunohistochemically, group-2 showed significant increases in IL-1β, TNF-α, MMP-13, and NOS-2 expression. However, all treatment groups exhibited reductions in these expression levels compared to group-2, with statistically significant decreases observed in groups-5 and 6 (P < 0.01).BA shows potential efficacy in reducing inflammation in experimental KOA model in rats. It may be a promising therapeutic agent for KOA, warranting further clinical studies for validation.

Remote ischemic preconditioning prevents sarcolemmal-associated proteolysis by MMP-2 inhibition.

The death of myocytes occurs through different pathways, but the rupture of the plasma membrane is the key point in the transition from reversible to irreversible injury. In the myocytes, three major groups of structural proteins that link the extracellular and intracellular milieus and confer structural stability to the cell membrane: the dystrophin-associated protein complex, the vinculin-integrin link, and the spectrin-based submembranous cytoskeleton. The objective was to determine if remote ischemic preconditioning (rIPC) preserves membrane-associated cytoskeletal proteins (dystrophin and β-dystroglycan) through the inhibition of metalloproteinase type 2 (MMP-2) activity. A second objective was to describe some of the intracellular signals of the rIPC, that modify mitochondrial function at the early reperfusion. Isolated rat hearts were subjected to 30min of global ischemia and 120min of reperfusion (I/R). rIPC was performed by 3 cycles of ischemia/reperfusion in the lower limb (rIPC). rIPC significantly decreased the infarct size, induced Akt/GSK-3 β phosphorylation and inhibition of the MPTP opening. rIPC improved mitochondrial function, increasing membrane potential, ATP production and respiratory control. I/R increased ONOO- production, which activates MMP-2. This enzyme degrades β-dystroglycan and dystrophin and collaborates to sarcolemmal disruption. rIPC attenuates the breakdown of β-dystroglycan and dystrophin through the inhibition of MMP-2 activity. Furthermore, we confirm that rIPC activates different intracellular pathway that involves the an Akt/Gsk3β and MPTP pore with preservation of mitochondrial function.

Metalloproteinase-2 in failed back surgery syndrome caused by epidural fibrosis: can it play a role in persistent pain?

Failed Back Surgery Syndrome (FBSS) occurs in 10-40% of patients treated surgically due to disk herniation (DH). There are several factors that can cause a predisposition to FBSS, but the exact pathomechanism has not been elucidated. The aim of this study was to investigate Metalloproteinase-2 (MMP-2) and Tissue Inhibitor of Metalloproteinase-2 (TIMP-2) activities in a homogeneous group of FBSS patients with epidural fibrosis in comparison to its activity in patients with surgically treated DH. DH, FBSS, and control (CG) groups consisted of 30 subjects. The patients were assessed clinically by the Numerical Rating Scale (NRS), McGill Pain Questionnaire (SF -MPQ), Oswestry Disability Index (ODI), and Beck Depression Inventory (BDI). Serum concentrations of MMP-2 and TIMP-2 were measured by using the immunoenzymatic method. There was a significantly higher MMP-2 expression (medians: 4797.49 vs. 2656.65; p < 0.0001) and TIMP-2 concentration (medians: 166.40 vs. 109.60; p < 0.0001) in the DH compared to the CG. Significantly higher MMP-2 expression (4219.95 vs. 2656.65; p < 0.0001) and TIMP-2 concentration (medians: 150.17 vs. 109.60; p = 0.0003) were also found in the FBSS compared to the CG. The activity of MMP-2, measured as MMP-2/TIMP-2, did not significantly change between the DH, FBSS, and CG. MMP2 expression (p < 0.0001) and TIMP-2 concentration (p < 0.0001) were significantly higher in the DH than FBSS. Results indicate the presence of a contribution of MMP-2 and TIMP-2 in DH and FBSS. Unchanged activity of MMP-2 can indicate an insufficiency in the MMP-2 repair system in both diseases. Lower MMP-2 expression and TIMP-2 concentration in the FBSS group can reflect the chronicity of the process.

Sexual dimorphism of the synovial transcriptome underpins greater PTOA disease severity in male mice following joint injury

Osteoarthritis is a disease with sex-dependent prevalence and severity in both humans and animal models. We sought to elucidate sex differences in synovitis, mechanical sensitization, structural damage, bone remodeling, and the synovial transcriptome in the anterior cruciate ligament rupture (ACLR) mouse model of post-traumatic osteoarthritis (PTOA). Male and female 12-week-old C57/BL6J mice were randomized to Sham or noninvasive ACLR with harvests at 7d or 28d post-ACLR (n=9 per sex in each group - Sham, 7d ACLR, 28d ACLR). Knee hyperalgesia, mechanical allodynia, and intra-articular MMP activity (via intravital imaging) were measured longitudinally. Trabecular and subchondral bone remodeling and osteophyte formation were assessed by µCT. Histological scoring of PTOA and synovitis and anti-MMP13 immunostaining were performed. NaV1.8-Cre;tdTomato mice were used to document localization and sprouting of nociceptors. Bulk RNA-seq of synovium in Sham, 7d, and 28d post-ACLR, and contralateral joints (n=6 per group per sex) assessed injury-induced and sex-dependent gene expression. Male mice exhibited more severe joint damage at 7d and 28d and more severe synovitis at 28d, accompanied by 19% greater MMP activity, 8% lower knee hyperalgesia threshold, and 43% lower hindpaw withdrawal threshold in injured limbs compared to female injured limbs. Females had injury-induced catabolic responses in trabecular and subchondral bone, whereas males exhibited 133% greater normalized osteophyte volume relative to females and sclerotic remodeling of trabecular and subchondral bone. NaV1.8+ nociceptor sprouting in subchondral bone and medial synovium was induced by injury and comparable between sexes. RNA-seq of synovium demonstrated similar injury-induced transcriptomic programs between the sexes at 7d, but only female mice exhibited a transcriptomic signature indicative of synovial inflammatory resolution by 28d, whereas males had persistent pro-inflammatory, pro-fibrotic, pro-neurogenic, and pro-angiogenic gene expression. Male mice exhibited more severe overall joint damage and pain behavior after ACLR, which was associated with persistent activation of synovial inflammatory, fibrotic, and neuroangiogenic processes, implicating persistent synovitis in driving sex differences in murine PTOA.

The essential role of the ERK activation in large T antigen of BK polyomavirus regulated cell migration

Recent studies have suggested that BK polyomavirus (BKPyV) may be associated with the development of urothelial carcinoma. In Merkel cell carcinoma, TAg and tAg are the major viral proteins of Merkel cell polyomavirus with oncogenic potential. In this study, we aimed to distinguish the role of TAg and tAg in cell migration. Our result demonstrated that ERK was phosphorylated in human renal tubular cells expressing its TAg and tAg after BKPyV infection. Treatment with the ERK inhibitor U0126 suppressed BKPyV gene expression and reduced BKPyV replication. Both TAg and tAg induced cell migration via ERK-dependent signaling. Furthermore, the expression of TAg and tAg had a significant regulatory effect on focal adhesion molecules in renal proximal tubular cells, which strongly suggests that alterations in the focal adhesion complexes are critically involved in TAg and tAg-induced cell migration. Gelatin zymography profiling revealed that TAg regulates the expression and activity of MMP-2 and MMP-9, but not tAg. Interestingly, TAg regulates the expression and activity of MMP-9 through ERK signaling, whereas MMP-2 is regulated through an ERK-independent pathway. Unbalanced ERK pathway activity is frequently observed in many cancers, while MMP proteins are usually overexpressed in aggressive tumors. These findings support the view that BKPyV is an oncogenic virus.

Hydroalcoholic Extract of Psoralea drupacea Inhibits Proliferation and Migration of Hepatocellular Carcinoma Cells and Decreases Angiogenesis in Chick Chorioallantoic Membrane

Objective: Hepatocellular carcinoma is one of the leading causes of cancer-related death worldwide. Experimental studies reported that some plants in the genus of Psoralea (Fabaceae family) show anticancer potential. The present study aimed to evaluate the effects of Psoralea drupacea extract (PDE) on HepG2 liver cancer cells. Methods: The proliferation, cell cycle, and migration of HepG2 cells were determined by thiazolyl blue tetrazolium bromide test, propidium iodide staining, and scratch assay, respectively. The effects of PDE on the activity of matrix metalloproteinases (MMPs) and angiogenesis were evaluated by the gelatin zymography method and chicken chorioallantoic membrane model, respectively. Results: The culture of HepG2 cells in the presence of PDE (24 hr and 48 hr) significantly reduced their viability (at a concentration of ≥ 50 µg/mL) and increased the percentage of cells in the sub-G1 stage. PDE also increased the antiproliferative and proapoptotic activities of doxorubicin (3 and 6 µg/mL). The extract significantly decreased the generation of reactive oxygen species and lipid peroxidation in the cells. Moreover, PDE (25 and 50 µg/mL) significantly suppressed the migration ability of HepG2 cells, which was associated with inhibition in the activity of MMP2 and MMP9 (50 µg/mL). Furthermore, treatment with PDE significantly reduced the number and diameter of vessels in the chick chorioallantoic membrane. Conclusion: PDE decreased the survival and cell cycle progression of liver cancer cells through a mechanism other than oxidative stress. This extract also showed an anti-angiogenesis effect and diminished the migration ability of HepG2 cells by inhibiting MMP activity

HDAC1 is Involved in Neuroinflammation and Blood-Brain Barrier Damage in Stroke Pathogenesis.

Stroke is a common cause of disability and mortality worldwide; however, effective therapy remains limited. In stroke pathogenesis, ischemia/reperfusion injury triggers gliosis and neuroinflammation that further activates matrix metalloproteinases (MMPs), thereby damaging the blood-brain barrier (BBB). Increased BBB permeability promotes macrophage infiltration and brain edema, thereby worsening behavioral outcomes and prognosis. Histone deacetylase 1 (HDAC1) is a repressor of epigenomic gene transcription and participates in DNA damage and cell cycle regulation. Although HDAC1 is deregulated after stroke and is involved in neuronal loss and DNA repair, its role in neuroinflammation and BBB damage remains unknown. The rats with cerebral ischemia were evaluated in behavioral outcomes, levels of inflammation in gliosis and cytokines, and BBB damage by using an endothelin-1-induced rat model with cerebral ischemia/reperfusion injury. The results revealed that HDAC1 dysfunction could promote BBB damage through the destruction of tight junction proteins, such as ZO-1 and occludin, after stroke in rats. HDAC1 inhibition also increased the levels of astrocyte and microglial gliosis, tumor necrosis factor-alpha, interleukin-1 beta, lactate dehydrogenase, and reactive oxygen species, further triggering MMP-2 and MMP-9 activity. Moreover, modified neurological severity scores for the cylinder test revealed that HDAC1 inhibition deteriorated behavioral outcomes in rats with cerebral ischemia. HDAC1 plays a crucial role in ischemia/reperfusion-induced neuroinflammation and BBB damage, thus indicating its potential as a therapeutic target.

S-nitrosylation of EMMPRIN influences the migration of HSCs and MMP activity in liver fibrosis.

The mechanism of extracellular matrix metalloproteinase inducer (EMMPRIN) in the regulation of liver fibrosis has not been clarified. This study aims to investigate the role of EMMPRIN S-nitrosylation (SNO) in the regulation of hepatic stellate cell (HSC) migration and matrix metalloproteinase (MMP) activities in liver fibrosis. The results from the tissue microarrays and rat/mouse liver tissues suggest that EMMPRIN mRNA and protein levels in the fibrotic livers are lower than those in the corresponding normal control livers, but higher SNO level of EMMPRIN in fibrotic liver area was shown by immunohistochemistry, immunofluorescence staining, and biotin-switch assay conversely in vivo. Primary EMMPRIN comes from hepatocytes and liver sinus epithelial cells (LSECs) rather than quiescent HSCs. To mimic the uptake of extrinsic EMMPRIN, supernatants from mouse primary hepatocytes/293 cells transfected with EMMPRIN wild-type plasmids (WT) and EMMPRIN SNO site (cysteine 87) mutation plasmids (MUT) were collected and added to JS-1/LX2 cell medium. The MUT EMMPRIN diminishes SNO successfully, enhances the activities of MMP2 and MMP9, and subsequently increases HSC migration. In conclusion, SNO of EMMPRIN influences HSC migration and MMP activities in liver fibrosis. This finding may shed light on the possible regulatory mechanism of MMPs in ECM accumulation in liver fibrosis.

Unlocking Drug Resistance in Multiple Myeloma: Adipocytes as Modulators of Treatment Response.

Multiple myeloma (MM) is an incurable hematological malignancy characterized by the clonal proliferation of malignant plasma cells. Despite the development of a diverse array of targeted drug therapies over the last decade, patients often relapse and develop refractory disease due to multidrug resistance. Obesity is a growing public health threat and a risk factor for multiple myeloma, although the mechanisms by which obesity contributes to MM growth and progression have not been fully elucidated. In the present study, we evaluated whether crosstalk between adipocytes and MM cells promoted drug resistance and whether this was amplified by obesity. Human adipose-derived stem cells (ASCs) from nineteen normal (BMI = 20-25 kg/m2), overweight (25-30 kg/m2), or obese (30-35 kg/m2) patients undergoing elective liposuction were utilized. Cells were differentiated into adipocytes, co-cultured with RPMI 8226 or U266B1 multiple myeloma cell lines, and treated with standard MM therapies, including bortezomib or a triple combination of bortezomib, dexamethasone, and lenalidomide. We found that adipocytes from overweight and obese individuals increased cell adhesion-mediated drug resistance (CAM-DR) survival signals in MM cells, and P-glycoprotein (P-gp) and multidrug resistance-associated protein (MRP) drug transporter expression. Further, co-culture enhanced in vitro angiogenesis, MMP-2 activity, and protected MM cells from drug-induced decreases in viability. In summary, we provide an underlying mechanism by which obesity can impair the drug response to MM and allow for recurrence and/or disease progression.

Extraction and purification of narirutin and hesperidin from green yuzu (Citrus junos) and evaluating their biological activities

This study examines how to obtain high levels of two major flavonoids, hesperidin and narirutin, from GY (green yuzu) powder and investigate their biological properties. We selected solvent extraction and enzyme treatment conditions before purification with an HP-20 Diaion column. We determined the appropriate extracted condition as 5 % GY with a 50 % ethanol solvent, and the enzyme treatment condition as 0.05 % RF enzyme for 24 h. Purification using 50 % ethanol as an eluent resulted in highly-enriched narirutin and hesperidin (50.78 % and 42.43 %, respectively). We also investigated the biological activities of extracts from each step, including the assessment of MMP-1 and tyrosinase inhibition activities, antibacterial properties, cytotoxicity, and anti-inflammatory activities. The extracts exhibited desirable properties such as skin elasticity, whitening effects, and anti-inflammatory effects in LPS-simulated RAW 264.7 cells. Highly purified extracts showed better antibacterial activity against food and dermatological pathogens. The results suggested that narirutin and hesperidin concentration and purity may influence the extracts’ biological effects. Overall, this study provides valuable insights into the improved extraction and purification of narirutin and hesperidin from GY powder and their biological activities, highlighting their potential as a functional food and health product ingredients.

Substrate Stiffness Modulates TGF-β Activation and ECM-Associated Gene Expression in Fibroblasts.

Transforming growth factor-β (TGF-β) is a multifunctional cytokine that regulates the expression of ECM-associated genes during early injury. Tissue fibrosis development is driven by synergistic cues between the evolving biochemical and mechanical milieu. Few studies have addressed the role of substrate stiffness on TGF-β activity and extracellular matrix (ECM)-associated genes. We used a commercial formulation of polydimethylsiloxane (PDMS) to fabricate substrates of 40 kPa, 300 kPa, and 1.5 MPa stiffness, and cultured the HMF3S fibroblasts on substrates. We quantified TGF-β protein secreted by HMF3S cells on different substrates using a TGF-β responsive promoter reporter assay. We also tested for variations in gene expression levels on the substrates using RT-PCR and Western blotting and determined the MMP-2 and MMP-9 activities with gelatin zymography. The results showed that TGF-β protein activation was significantly compromised at lower stiffnesses. The expression of integrin α5 decreased on lower stiffness substrates and correlated with inefficient TGF-β protein activation. Collagen I, collagen III, and MMP-2 expression levels were lower on softer substrates; there was little MMP-9 activity on all substrates. Cell and nuclear morphologies were more rounded on compliant substrates, correlating with increased tubulin expression. Proliferations were higher on stiffer substrates, whereas cells on softer substrates showed cell cycle arrest. These results demonstrated critical feedback mechanisms between substrate stiffness and ECM regulation by fibroblasts, relevant in fibrosis.

Type I collagen proteolysis by matrix metalloproteinase-2 contributes to focal adhesion kinase activation and vascular smooth muscle cell proliferation in the aorta in early hypertension

IntroductionIncreased matrix metalloproteinase (MMP)-2 activity contributes to increase vascular smooth muscle cell (VSMC) proliferation in the aorta in early hypertension by cleaving many proteins of the extracellular matrix. Cleaved products from type I collagen may activate focal adhesion kinases (FAK) that trigger migration and proliferation signals in VSMC. We therefore hypothesized that increased activity of MMP-2 proteolyzes type I collagen in aortas of hypertensive rats, and thereby, induces FAK activation, thus leading to increased VSMC proliferation and hypertrophic remodeling in early hypertension. MethodsMale Sprague-Dawley rats were submitted to renovascular hypertension by the two kidney-one clip (2K1C) model and treated with doxycycline (30 mg/kg/day) by gavage from the third to seventh-day post-surgery. Controls were submitted to sham surgery. Systolic blood pressure (SBP) was measured daily by tail-cuff plethysmography and the aortas were processed for zymography and Western blot for MMP-2, pFAK/FAK, integrins and type I collagen. Mass spectrometry, morphological analysis and Ki67 immunofluorescence were also done to identify collagen changes and VSMC proliferation. A7r5 cells were stimulated with collagen and treated with the MMP inhibitors (doxycycline or ARP-100), and with the FAK inhibitor PND1186 for 24 h. Cells were lysed and evaluated by Western blot for pFAK/FAK. Results2K1C rats developed elevated SBP in the first week as well as increased expression and activity of MMP-2 in the aorta (p < 0.05 vs. Sham). Treatment with doxycycline reduced both MMP activity and type I collagen proteolysis in aortas of 2K1C rats (p < 0.05). Increased pFAK/FAK and increased VSMC proliferation (p < 0.05 vs. Sham groups) were also seen in the aortas of 2K1C and doxycycline decreased both parameters (p < 0.05). Higher proliferation of VSMC contributed to hypertrophic remodeling as seen by increased media/lumen ratio and cross sectional area (p < 0.05 vs Sham groups). In cell culture, MMP-2 cleaves collagen, an effect reversed by MMP inhibitors (p < 0.05). Increased levels of pFAK/FAK were observed when collagen was added in the culture medium (p < 0.05 vs control) and MMP and FAK inhibitors reduced this effect. ConclusionsIncrease in MMP-2 activity proteolyzes type I collagen in the aortas of 2K1C rats and contributes to activate FAK and induces VSMC proliferation during the initial phase of hypertension.

Detection and activity of MMP-2 and MMP-9 in Leishmania amazonensis and Leishmania braziliensis promastigotes

Metalloproteinases (MMPs) are remarkable zinc-dependent endopeptidases, critical for degrading components of the extracellular matrix, thus actively influencing cell migration. Their impact on intracellular parasites, such as the enigmatic protozoan Leishmania, elicits intriguing queries. This study explores into the untapped territory of MMP-2 and MMP-9 within Leishmania spp. promastigotes. Notably, we successfully detected and quantified these MMPs, while also evaluating their activity in two distinct Leishmania species—L. amazonensis (La) and L. braziliensis (Lb)—at various growth stages and isolated from distinct clinical tegumentar disease forms. The results unveiled the presence of MMP-2 and MMP-9 in both species, albeit with distinct localization patterns. Specifically, MMP-9 exhibited significantly higher gelatinolytic activity in La when compared to Lb. Moreover, our data cleverly illustrated the presence and release of MMP-2 and MMP-9 by La and Lb promastigotes, exposing their ability to invade and migrate within a collagen matrix. This pioneering study establishes a compelling correlation between MMP-2 and MMP-9 and their potential role in the dynamics of La and Lb infection. Suggesting their potential as prognostic markers for severe leishmaniasis and promising target molecules for therapeutic interventions, this research opens new avenues for combatting this debilitating parasitic disease.