Impact of low-level laser therapy as an adjunct to non-surgical periodontal treatment on the levels of tissue plasminogen activator and plasminogen activator inhibitor 1 in Stage 3-4, Grade C periodontitis patients: a split-mouth, randomized control study.

AIM: To investigate the effects of low-level laser therapy (LLLT) as an adjunct to non-surgical periodontal treatment (NSPT) on the plasminogen-activating system. MATERIALS AND METHODS: Stage 3-4 Grade C periodontitis and age-gender-matched healthy individuals participated in the split-mouth study (ClinicalTrials.gov identifier, NCT05233501). The study groups were Periodontitis/NSPT (Sham); Periodontitis/NSPT + LLLT (LLLT); Healthy (Control). Following NSPT, LLLT was applied on Days 0, 2 and 7. Clinical parameters were recorded at baseline and on Day 30. Gingival crevicular fluid (GCF) was collected at baseline, on days 7, 14, and 30; tissue-type plasminogen activator (tPA) and plasminogen activator inhibitor-1 (PAI-1) levels were measured with ELISA. RESULTS: Clinical parameters, total GCF tPA (tPAt) and PAI-1 (PAI-1t) levels significantly reduced in LLLT and Sham groups (< 0.001). GCF tPAt levels in LLLT were significantly lower (< 0.05) than Sham on Day 7. GCF tPAt levels in periodontitis groups were significantly higher than the Control at baseline, on Days 7 and 14 (< 0.01). By Day 30, both groups decreased to control levels (> 0.05). GCF PAI-1t levels were significantly lower in LLLT than the Sham on day 30 (< 0.01), comparable to healthy controls (> 0.05). CONCLUSION: Adjunctive LLLT modulates the plasminogen activating system in severe periodontitis by altering GCF tPA and PAI-1 levels. CLINICAL RELEVANCE: LLLT as an adjunct to non-surgical periodontal treatment in patients with Stage 3-4 Grade C leads to reduced plasminogen activation.

The effect of low-level laser therapy as an adjunct to non-surgical periodontal treatment on gingival crevicular fluid levels of transforming growth factor-beta 1, tissue plasminogen activator and plasminogen activator inhibitor 1 in smoking and non-smoking chronic periodontitis patients: A split-mouth, randomized control study.

BACKGROUND AND OBJECTIVE: This study aimed to investigate the effects of low-level laser therapy (LLLT) as an adjunct to scaling and root planing (SRP) on smoking and non-smoking patients with chronic periodontitis. MATERIAL AND METHODS: The study was conducted using a split-mouth design with 30 patients with chronic periodontitis (15 smokers, 15 non-smokers) and 30 healthy individuals matched for age, sex and smoking status as controls. Groups were constituted as follows: Cp+SRP+Sham: non-smokers with chronic periodontitis treated with SRP; Cp+SRP+LLLT: non-smokers with chronic periodontitis treated with SRP+LLLT; SCp+SRP+Sham: smokers with chronic periodontitis treated with SRP; SCp+SRP+LLLT: smokers with chronic periodontitis treated with SRP+LLLT; C: control group comprised of periodontally healthy non-smokers; SC: control group comprised of periodontally healthy smokers. LLLT was first applied on the same day as SRP and again on days 2 and 7 after SRP treatment. Clinical parameters were recorded before non-surgical periodontal treatment (baseline) and on day 30. Gingival crevicular fluid samples were collected before periodontal treatment (baseline) and during follow-up visits on days 7, 14 and 30. Gingival crevicular fluid transforming growth factor (TGF)-ß1, tissue plasminogen activator (tPA) and plasminogen activator inhibitor 1 (PAI-1) levels were measured using enzyme-linked immunosorbent assay. RESULTS: All clinical parameters showed significant reductions between baseline and day 30 following SRP treatment in both the LLLT and sham groups (P<.001). No significant differences were observed between the LLLT and sham groups of either the smokers or non-smokers (P>.05). Gingival crevicular fluid PAI-1 levels decreased significantly in the SCp+SRP+sham and SCp+SRP+LLLT groups (P<.05), and gingival crevicular fluid tPA levels decreased significantly in the Cp+SRP+sham, Cp+SRP+LLLT and SCp+SRP+LLLT groups (P<.05). Gingival crevicular fluid TGF-ß1 levels decreased significantly in all treatment groups (P<.05). Although no significant differences were found between the gingival crevicular fluid PAI-1, tPA and TGF-ß1 levels of the LLLT versus sham groups (P>.05) at any of the time points measured, both LLLT groups showed significant reductions in tPA/PAI-1 ratios over time. CONCLUSION: Within the limits of this study, LLLT may be understood to play a role in the modulation of periodontal tissue tPA and PAI-1 gingival crevicular fluid levels, particularly in smoking patients with chronic periodontitis, and may thus be recommended as an adjunct to non-surgical periodontal treatment.

Up-Regulation of PAI-1 and Down-Regulation of uPA Are Involved in Suppression of Invasiveness and Motility of Hepatocellular Carcinoma Cells by a Natural Compound Berberine.

Hepatocellular carcinoma (HCC) is the second leading cause of cancer-related death and its prognosis remains poor due to the high risk of tumor recurrence and metastasis. Berberine (BBR) is a natural compound derived from some medicinal plants, and accumulating evidence has shown its potent anti-tumor activity with diverse action on tumor cells, including inducing cancer cell death and blocking cell cycle and migration. Molecular targets of berberine involved in its inhibitory effect on the invasiveness remains not yet clear. In this study, we identified that berberine exhibits a potent inhibition on the invasion and migration of HCC cells. This was accompanied by a dose-dependent down-regulation of expression of Cyclooxygenase-2 (COX-2), nuclear factor kappa B (NF-κB), urokinase-type plasminogen activator (uPA) and matrix metalloproteinase (MMP)-9 in berberine-treated HCC cells. Furthermore, berberine inactivated p38 and Erk1/2 signaling pathway in HCC cells. Primarily, this may be attributed to the up-regulation of plasminogen activator inhibitor-1 (PAI-1), a tumor suppressor that can antagonize uPA receptor and down-regulation of uPA. Blockade of uPA receptor-associated pathways leads to reduced invasiveness and motility of berberine-treated HCC cells. In conclusion, our findings identified for the first time that inactivation of uPA receptor by up-regulation of PAI-1 and down-regulation of uPA is involved in the inhibitory effect of berberine on HCC cell invasion and migration.

Activation of fibrinolysis by reinfusion of unwashed salvaged blood after total knee arthroplasty.

In 19 patients with RA and 20 with OA who underwent autotransfusion with unwashed salvaged blood (USB) after total knee arthroplasty, we performed serial measurement of D-dimer, FgDP, t-PA, and PIC in the plasma and salvaged blood. The PIC level at the completion of salvaged blood transfusion was closely correlated with the volume of USB reinfused in the RA group (p<0.001). The t-PA level of salvaged blood showed a significant positive correlation with the total volume of postoperative drainage in both the RA and OA groups (p<0.05). The increase of total postoperative drainage associated with reinfusion of USB is largely caused by an increase of t-PA in the salvaged blood.

A citrus extract containing flavanones represses plasminogen activator inhibitor-1 (PAI-1) expression and regulates multiple inflammatory, tissue repair, and fibrosis genes in human colon fibroblasts.

The consumption of flavonoid-rich Citrus extracts has been associated with multiple beneficial effects including anti-inflammatory properties, but the potential effects on the inflammatory responses in the gut have not been thoroughly investigated. We used microarrays to search for molecular changes induced in human colon fibroblasts in response to the exposure to a flavanone-rich bitter orange extract under physiologically representative conditions. Dietary nontoxic levels of the predigested extract induced moderate but significant changes in the expression of genes associated with tissue repair and inflammation. Among the top regulated genes, plasminogen activator inhibitor 1 (PAI-1) was downregulated, and the matrix metallopeptidase 12 (MMP-12) was upregulated (mRNA and protein levels). Both proteins are involved in extracellular matrix (ECM) remodeling and fibroblast migration. The extract also affected the fibroblast migration and reduced monocyte adhesion, but the response was different in unstimulated cells and in cells pretreated with TNF-alpha. Collectively, these results were indicative of a moderate activation of the colon fibroblast inflammation-related function after exposure to the extract. Further investigations are required to identify the in vivo role of this Citrus derived extract in the maintenance of the normal balance in the intestine and in the pathogenesis of inflammatory diseases.

[Influence of tea polyphenols on the damage of fibrinolytic system induced by homocysteine in human umbilical vein endothelial cells].

OBJECTIVE: To study the influence of tea polyphenols (TP) on cells form and fibrinolytic system which focused on tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor-1 (PAI-1) induced by homocysteine (Hcy) in human umbilical vein endothelial cells (HUVEC). METHODS: HUVEC were divided into two groups, one was administered with TP and Hey at the same time, the other was firstly treated with Hey for 12 hours and followed by administering TP. Each group was divided into some groups depending on different concentrations of TP. The t-PA and PAI-1 levels of HUVEC supernatant were determined by ELISA assay. RESULTS: Compared with the groups that just treated with Hcy, medium concentration (4 and 8 microg/ml) and high concentration (16 microg/ml) of TP could strengthen cells stereoscopic effect and decrease endochylema particles; the results of ELISA demonstrated that the concentrations of PAI-1 were increased in the groups that just treated with Hey compared with control group (P < 0.05), administrations of medium and high dose of TP could decrease the concentration of PAI-1, and the concentration of PAI-1 were decreased depending on the increase of TP concentration (P < 0.05), there was no difference between low concentration TP group and the group only treated with Hcy. There were no differences on the concentrations of t-PA among each groups. CONCLUSION: The proper concentration of TP could restore the damage of HUVEC form and decrease the concentration of PAI-1 induced by Hey.

Emerging biomarkers and new understanding of traditional markers in personalized therapy for breast cancer.

The era of personalized medicine is likely to see an escalation in the use of biomarkers to ensure breast cancer patients receive optimal treatment. A combination of prognostic and predictive biomarkers should enable better quantification of the residual risk faced by patients and indicate the potential value of additional treatment. Established biomarkers such as estrogen receptor and progesterone receptor already play a significant role in the selection of patients for endocrine therapy. Human epidermal growth factor receptor 2 (HER2) is recognized as a strong predictor of response to trastuzumab whereas, more recently, the role of estrogen receptor and HER2 as negative and positive indicators for chemotherapy has also been explored. Ki67 has traditionally been recognized as a modest prognostic factor, but recent neoadjuvant studies suggest that on-treatment measurement may be a more effective predictor of treatment efficacy for both endocrine treatment and chemotherapy. The last decade has seen the emergence of numerous multigene expression profiles that aim to outdo traditional predictive and prognostic factors. The Oncotype DX assay and the MammaPrint profile are currently undergoing prospective clinical trials to clearly define their role. Other gene expression-based assays also show potential but are yet to be tested clinically. Rigorous comparison of these emerging markers with current treatment selection criteria will be required to determine whether they offer significant benefit to justify their use.

Can the aggregation be a new approach for understanding the mechanism of Traditional Chinese Medicine?

"Frequent hitter" phenomenon emerged in the high-throughput screening; one of the most common mechanisms behind artifactual inhibition is that some organic molecules formed large colloid-like aggregates which are able to sequester and thereby inhibit enzymes. To investigate the situation in Traditional Chinese Medicine (TCM), 60 medicinal herbs and 24 Chinese herbal formulae were detected by dynamic light scattering (DLS), and aggregates were observed in all the 84 solution mixtures. The aggregates of two Chinese herbal formulae, 'Xue-Fu-Zhu-Yu Tang' (XF) and 'Jing-Guan Tang' (JG), were not only able to survive in the gastro-intestinal environment, but also had the ability to pass through the monolayer of the Caco-2 cell. The activities of XF and JG against three cardiovascular targets were also aggregates-related. Based on these findings, a new possible mechanism of the action of Chinese medicine was proposed.

Vitamin C and E supplementation in women at risk of preeclampsia is associated with changes in indices of oxidative stress and placental function.

OBJECTIVE: We have previously reported a reduced incidence of preeclampsia in women who were at risk and were taking vitamin C (1000 mg/d) and vitamin E (400 IU/d) supplements. In this study, we determined whether supplementation in the same cohort was associated with an improvement in indices of placental dysfunction and oxidative stress toward values determined in women who were at low risk of preeclampsia. STUDY DESIGN: Seventy-nine women who were at high risk and who were taking vitamin supplements and 81 who were taking placebos were compared with 32 women who were at low risk and who were not taking supplements who were studied simultaneously. RESULTS: Indices of oxidative stress and placental function were abnormal in the placebo group. When the placebo group was compared with the women who were at low risk, ascorbic acid, plasminogen activator inhibitor-2, and placenta growth factor concentrations were decreased; and 8-epi-prostaglandin F(2alpha),leptin, and the plasminogen activator inhibitor-1/-2 ratio were increased. In the group that received vitamin supplements, ascorbic acid, 8-epi-prostaglandin F(2alpha), leptin, and plasminogen activator inhibitor-1/-2 values were similar to women who were at low risk. CONCLUSION: Antioxidant supplementation in women who were at risk of preeclampsia was associated with improvement in biochemical indices of the disease.

Vitamins C and E attenuate plasminogen activator inhibitor-1 (PAI-1) expression in a hypercholesterolemic porcine model of angioplasty.

BACKGROUND: The plasminogen activator inhibitor-1 (PAI-1), which modulates fibrinolysis and cell migration, may influence proteolysis and neointimal formation in the arterial wall contributing to restenosis after vascular injury. Antioxidants have been proposed as inhibiting multiple proatherogenic events. We explore the effect of vitamins C and E on PAI-1 expression in an experimental model of angioplasty in hypercholesterolemic pigs. METHODS AND RESULTS: A total of 44 Yucatan minipigs were divided into three diet groups: a normal-cholesterol (NC), a high-cholesterol (HC), and a high-cholesterol plus vitamins C+E (HCV) group. Balloon injury was induced in the right internal iliac artery 4 weeks after initiation of either dietary regimen, and plasma and tissue samples were taken at different time periods to measure PAI-1 activity and vascular inhibitor expression. The cholesterol-rich diet induced an increased in vascular PAI-1 expression in the intima, media and adventitia which was markedly reduced in the HCV group. After injury, severe structural changes were observed in NC and HC animals associated with increased systemic PAI-1 activity (P<0.001) and local PAI-1 expression being more intense in HC group. Vitamins C and E significantly reduced plasma PAI-1 activity (P=0.018) and attenuated the inhibitor expression as compared with HC. CONCLUSIONS: This experimental study in a porcine model of hypercholesterolemia demonstrates that vitamins C and E reduce local and systemic PAI-1 induced after angioplasty as well as the hypercholesterolemia-induced vascular PAI-1.

Prevention of renal fibrin deposition in endotoxin-induced DIC through inhibition of PAI-1.

Plasminogen activator inhibitor-1 (PAI-1) increases in endotoxemia thus possibly cooperating in altering the hemostatic balance in a prothrombotic direction. The effect of the inhibition of PAI-1 with the monoclonal antibody MA-33B8 was studied systemically and in kidneys in a lapine model of endotoxin-induced disseminated intravascular coagulation (DIC). The increase in plasmatic PAI activity in the control group (n = 9) was inhibited in the MA-33B8 treated rabbits (n = 5). Control rabbits showed renal fibrin deposits, whereas only one of the MA-33B8 rabbits did so. These results were confirmed immunohistochemically in kidneys as PAI-1 immunostaining was seen inside the glomeruli and larger vessels in the control group, whereas MA-33B8 rabbits showed a remarkable decrease, demonstrating that MA-33B8 successfully inhibited PAI-1 in the kidneys as well. Therefore evidence for the important role of PAI-1 in fibrin generation in endotoxin-induced DIC is presented, suggesting that strategies aiming at its reduction can be useful in this pathology.

The effects of six months of treatment with a low-dose of conjugated oestrogens in menopausal women.

OBJECTIVE: Hormone replacement therapy (HRT) is usually prescribed as medium- to high-dose formulations. Little is known, however, about dose-dependency of oestrogen effects on plasma hormone levels, markers of cardiovascular risk in lipid metabolism and the haemostatic system, or markers of bone turnover. SUBJECTS AND DESIGN: In an open trial, three groups of 12 or 13 healthy, non-obese postmenopausal women received conjugated equine oestrogens (CEE) for 6 months at doses of 0.3 mg/day (group 1), 0.6 mg/day (group 2) or 1.25 mg/day (group 3). From day 1 to day 10, CEE was administered alone, and from day 11 to day 21, in combination with 5 mg of medrogestone. Each treatment cycle was followed by a pause of 7 days. Fasting blood samples were obtained before treatment as well as on days 10, 21 and 28 of the first, third and sixth months on treatment. All results obtained on day 10 were grouped together as phase A, on day 21 as phase B, and on day 28 as phase C. MEASUREMENTS: Plasma concentrations of oestradiol (E), dehydroepiandrosterone sulphate (DHEA-S), total testosterone (T), FSH, PRL, sex hormone binding globulin (SHBG), type I procollagen propeptide (PICP) and the cross-linked carboxyterminal telopeptide of type I collagen (ICTP), total cholesterol, HDL-cholesterol, triglycerides (TG), apolipoprotein (apo) A-1, apo B, lipoprotein(a)[Lp (a)], fibrinogen, factor VIIc and plasminogen activator inhibitor 1 (PAI-1) were evaluated with commercially available kits. RESULTS: Dose-dependently, the three regimens increased E, SHBG and factor VIIc activity and decreased FSH, DHEAS, cholesterol, LDL-cholesterol and apoB. HDL-cholesterol and apoA-1 were slightly decreased in group 1 but increased in groups 2 and 3. The high CEE dosage in group 3 resulted in a significant increase of TG and decrease of Lp(a) and PAI-1. Markers of bone turnover were not significantly changed by any CEE dosage. CONCLUSIONS: Six months of treatment with 0.3 mg/day of conjugated equine oestrogen significantly lowers serum levels of total cholesterol and LDL-cholesterol without causing the adverse increases of triglycerides or factor VIIc, which were observed at higher doses. However, this low-dose treatment did not yield the maximal LDL-cholesterol lowering effect. Moreover, the positive effects of HRT on HDL-cholesterol, apolipoprotein A-I, lipoprotein (a) and plasminogen activator inhibitor-1 required at least the medium dose of 0.6 mg conjugated equine oestrogens per day. Therefore, further studies are needed to determine which dose of conjugated equine oestrogens has the optimal effect on cardiovascular risk and bone turnover.

Plasminogen activator inhibitor-1 (PAI-1) is not related to response to neoadjuvant chemotherapy in breast cancer.

There is no information available on the relation between response to chemotherapy and the high-risk phenotype assessed by uPA and/or PAI-1. The clinical situation of neoadjuvant chemotherapy provides a means of rapidly assessing the sensitivity of the primary tumour to cytotoxic drug regimens. The goal of the study was to assess prospectively the predictive value of PAI-1 for response to first-line chemotherapy. PAI-1 concentration was measured on hypertonic cytosolic extracts (0.4 M potassium chloride) by ELISA before chemotherapy on a drill biopsy sample of the tumour in 69 T2 and T3 breast cancer patients (median age 46 years). Oestrogen receptor (ER) (51% ER+), progesterone receptor (PR) (58% PR+), S-phase (median 4.0%) and ploidy were also assessed in the majority of cases. The clinical response to treatment was evaluated after four cycles of FAC or FEC regimen (5-fluorouracil, epidoxorubicin or doxorubicin and cyclophosphamide) (one cycle every 4th week). PAI-1 could be assayed in 29 post-chemotherapy surgical samples. The objective response rate (complete response plus partial response) was 59% (41 out of 69). PAI-1 expressed as gram of tissue (range 19-2370 ng g(-1) tissue) was highly correlated (r = 0.98) to PAI-1 expressed as mg protein (range 0.5-68 ng mg(-1) protein). No correlation between PAI-1 level and response could be observed, with any cut-off. The post- and pre-chemotherapy PAI-1 levels were correlated (r = 0.66). Of all biological parameters, only high S-phase (cut-off 5%) was slightly correlated (chi2 = 3.91, P = 0.05) to response. These data suggest that PAI-1 is not a predictive marker of response to chemotherapy in breast cancer and that its level is not altered by neoadjuvant chemotherapy.

The plasminogen-activating system in hepatic stellate cells.

Urokinase plasminogen activator (uPA) generates plasmin, a process inhibited by plasminogen-activator inhibitor (PAI)-1 and localized to the cell surface by binding of uPA to a specific receptor. Plasmin degrades extracellular matrix (ECM) both directly and by activation of matrix metalloproteinases (MMPs). Because stellate cells play a central role in the pathogenesis of liver fibrosis both via production of ECM proteins and through secretion of MMPs, their contribution to plasmin generation was assessed. Stellate cells were prepared from rat liver and cultured on plastic. Northern analysis showed cellular expression of messenger RNA (mRNA) for PAI-1, uPA, and uPA receptor. Zymography/reverse zymography identified cell-surface-associated uPA activity and uPA and PAI-1 in culture media. Net uPA activity in culture media was maximal after 7 days in culture and then declined, whereas PAI-1 antigen levels remained consistently elevated between 7 and 21 days in culture. Stellate cell-mediated plasmin generation was also seen in in vitro cultures supplemented with plasminogen. Because hepatic stellate cells (HSCs) contain retinoids and release them on activation, the effect of retinoic acid on the plasminogen-activating system was also assessed. Treatment of cultured HSCs with retinoic acid (1 micromol/L) increased uPA secretion 2.6-fold but did not alter PAI-1. We conclude that stellate cells synthesize key components of the plasminogen-activating system and generate plasmin and therefore have the ability to regulate MMP activation. Upregulation of uPA synthesis by retinoic acid may have implications in matrix remodeling in sites of stellate cell activation in which high concentrations of retinoids may be achieved.

Autologous blood transfusion in hip replacement. No effect on blood loss but less increase of plasminogen activator inhibitor in a randomized series of 80 patients.

80 patients underwent total hip replacement (THR) for primary coxarthrosis. In a randomized study, half of them donated 2 units of blood before operation. One unit was collected 4 weeks and one 2 weeks before the scheduled THR. All except 1 patient tolerated the predonations well. Total blood losses were similar in both groups. Additional bank blood was given in 7/38 in the predonation group, compared to 29/40 in the control group. Hemostatic parameters were studied in 10 consecutive patients in each group. Plasminogen activator inhibitor 1 (PAI-1), a possible risk parameter for thromboembolism, was significantly more increased postoperatively in the control group, which received only homologous blood. Platelet count, prothrombin complex, antithrombin III and von Willebrand factor antigen were significantly reduced and C reactive protein increased after surgery in both groups. We recommend predonation of 2 autologous units before a primary THR. In most cases, such predonation makes homologous blood transfusion unnecessary. The use of predonated blood causes no reduction of blood loss in THRs, but the increase in PAI-1 seen after homologous transfusions is avoided.

The acute effect of a single very high dose of N-3 fatty acids on coagulation and fibrinolysis.

The objective of the study was to investigate the acute effect of a single very high dose of n-3 PUFA on coagulation and fibrinolysis. Forty healthy volunteers were randomized into two groups to receive either 20 grams of n-3 PUFA or 20 grams of n-6 PUFA as a single dose at 6 p.m. with their evening meal. Coagulation and fibrinolysis were evaluated in the fasting state at 8 a.m. the next morning and compared to values obtained at 8 a.m. the day before, when the participants were on their habitual diets. PAI-1 activity in plasma increased by a mean of 62% in subjects randomized to receive n-3 PUFA despite that no changes could be demonstrated in t-PA antigen levels. PAI-1 activity was unaltered in the 20 controls receiving n-6 PUFA. Plasma fibrinogen, coagulation factor VII, thrombin-antithrombin complexes and D-dimer did not significantly change after either supplement. The substantial increase in levels of PAI-1 activity in plasma after a single very high dose of n-3 PUFA may limit the usefulness of single very high doses of n-3 PUFA in acute clinical conditions.