Levels Of TF Research Articles

Integrated network pharmacology and transcriptomics to explore the mechanism of compound Dihuang granule (CDG) protects dopaminergic neurons by regulating the Nrf2/HMOX1 pathway in the 6-OHDA/MPP+-induced model of Parkinson’s disease

BackgroundParkinson's disease (PD) is a degenerative neurological disease that worsens over time. Ferroptosis has been proven to contribute to PD pathogenesis. CDG exhibits neuroprotective effects. However, CDG's potential mechanism in PD therapy remains uncertain.PurposeThe purpose of this investigation is to ascertain the specific molecular mechanisms of CDG against neuronal ferroptosis and present an alternative option for PD management.MethodsNetwork pharmacology along with LC–MS were used to identify possible targets and candidate pathways. Then RNA-sequencing combined in the in vitro and in vivo experiments were utilized to validate these findings.ResultsAccording to network pharmacology prediction, NFE2L2, HMOX1 and PTGS2 may be the key genes for ferroptosis in PD. In the in vivo experiments, CDG ultimately improved the neurobehavior of PD rats by alleviating the damage of dopamine neurons, decreasing the levels of MDA, ROS and Fe2+, increasing the GSH level, inhibiting ferroptosis by decreasing ACSL4, TF, and PTGS2 expression levels, and increasing the GPX4, FTH, Nrf2, and HMOX1 levels. RNA-seq analysis showed the differential genes in Model and CDG group were all enriched in Nrf2 and HMOX1, and the enrichment analysis of these differential genes showed they were closely related to the ferroptosis. Subsequently, in vitro experiments, the CDG, OE-Nrf2 and OE-HMOX1 group showed more active cell vitality, with decreasing levels of MDA, ROS, Fe2+, ACSL4, TF and PTGS2, and increasing level GSH, GPX4, FTH, Nrf2 and HMOX1.ConclusionCDG has a neuroprotective involvement in alleviating ferroptosis by regulating the Nrf2/HMOX1 pathway. Moreover, this research offers pharmacological evidence supporting the applications of CDG for treating PD.

DEHP regulates ferritinophagy to promote testicular ferroptosis via suppressing SIRT1/PGC-1α pathway

To increase elasticity and flexibility, di-2-ethylhexyl phthalate (DEHP) is used in a variety of industrial products, but excessive exposure to it can pose a threat to human health. In epidemiological studies of population exposure to DEHP, attention has been paid to damage to the male reproductive system. However, the toxicological mechanism of DEHP regarding testicular injury is not well understood. We used Western blot analysis, transmission electron microscopy, fluorescence staining, transient transfection and assay kit to detect relevant indicators, and the results were as follows: After DEHP exposure, the expression levels of ACSL4, COX2, TF, FTH1, LC3, AMPK, p-AMPK, ULK1, p-ULK1, serum iron, tissue iron and MDA in the exposure group were significantly increased. The expression levels of GPX4, NCOA4, p62, SIRT1, and PGC-1α, as well as the contents of GSH and ATP, decreased. Electron microscopy showed that more autophagosomes were observed. Our findings suggest that exposure to DEHP induced ferritinophagy and ferroptosis in the testis. In vitro, the promoting effect of ferritinophagy on ferroptosis was verified by applying the autophagy inhibitor (3-MA) and si-NCOA4. Moreover, Mono-(2-ethylhexyl) phthalate (MEHP) inhibited the mitochondrial regulatory protein SIRT1/PGC-1α, leading to mitochondrial dysfunction. Changes in mitochondrial reactive oxygen species (MtROS) and energy over-activated AMPK/ULK1 autophagy pathway, and then promoted ferritinophagy, which increased the sensitivity of TM4 cells to ferroptosis. This research offers a theoretical framework for the prevention and management of DEHP-induced harm.

The association between periodontal microbial biomarkers and primary therapy outcome

ObjectiveThis study aims to analyse the association between the baseline microbial load of selected periodontopathogenic bacteria collected from gingival crevicular fluid (GCF) and the primary outcome of steps I and II therapy.Materials and methods222 patients with stage III periodontitis were included into this retrospective analysis that received steps 1 and 2 periodontal therapy without adjunctive systemic antibiotics. Baseline GCF samples were quantitatively analysed using ELISA-based kits for levels of periodontopathogens (Porphyromonas gingivalis (Pg), Aggregatibacter actinomycetemcomitans (Aa), Prevotella intermedia (Pi), Fusobacterium nucleatum (Fn), Treponema denticola (Td), and Tannerella forsythia (Tf)) and associated with the primary therapy outcome using a “treat-to-target” therapy endpoint (TE) defined as ≤ 4 sites with PD ≥ 5 mm six months after therapy.Results38.2% of the patients achieved TE. Patients failing to achieve TE revealed significantly increased levels of Pg, Fn, and Tf at baseline (Pg: p = 0.010, Fn: p = 0.008 Tf: p = 0.004). Multivariate binary logistic regression adjusted for sex, mean probing depth, diabetes, and current smoking status showed an independent relationship between Tf and the TE (aOR 2.570, p = 0.023).ConclusionIncreased microbial load is associated with decreased responsiveness to therapy. The findings suggest that specifically baseline Tf levels are associated with poorer treatment outcomes and might improve the accuracy of periodontal diagnosis.Clinical relevanceThe findings of this study support the concept of a critical biomass that is sufficient to induce and maintain an immune response within the periodontal pocket, which ultimately leads to irreversible tissue destruction. However, calculating this level in advance may serve as an early indicator for intervention.Key findingBaseline Tannerella forsythia levels are associated with poorer treatment outcome.

DNP and ATP modulate the pulp softening and breakdown in fresh longan by acting on the antioxidant system and the metabolisms of membrane lipids and cell wall polysaccharides

Compared to the control longan, DNP treatment elevated pulp breakdown index, reduced the values of pulp firmness, CSP, ISP, cellulose, and hemicellulose by enhancing the activities of PE, PG, Cx, XET, and β-Gal. Additionally, DNP treatment increased the levels of PLD, lipase, LOX, PA, and SFA, and decreased the values of PC, PI, USFA, U/S, and IUFA, displaying higher cell membrane permeability and more severe cell membrane damage in longan pulp. Furthermore, DNP treatment weakened the levels of SOD, CAT, APX, AsA, GSH, TP, and TF, thereby exacerbating ROS outbreak and MDA production. These results indicate that DNP treatment destroyed the antioxidant system to cause ROS eruption. This disruption further disturbed the metabolisms of membrane lipids and cell wall polysaccharides, leading to the breakdown of cell membrane and cell wall, and eventually aggravated longan pulp softening and breakdown. However, ATP treatment exhibited the opposite effects of DNP treatment.

The TF/Nrf2/GSTP1 pathway is involved in stress-induced hepatocellular injury through ferroptosis.

Stress triggers a comprehensive pathophysiological cascade in organisms. However, there is a substantial gap in the research regarding the effects of stress on liver function. This study aimed to investigate the impact of restraint stress on hepatocellular damage and elucidate the underlying molecular mechanisms. An effective mouse restraint stress model was successfully developed, and liver function analysis was performed using laser speckle imaging, metabolomics and serum testing. Alterations in hepatocyte morphology were assessed using haematoxylin and eosin staining and transmission electron microscopy. Oxidative stress in hepatocytes was assessed using lipid reactive oxygen species and malondialdehyde. The methylation status and expression of GSTP1 were analysed using DNA sequencing and,real-time PCR, and the expression levels of GPX4, TF and Nrf2 were evaluated using real-time quantitative PCR, western blotting, and immunohistochemical staining. A stress-induced model was established invitro by using dexamethasone-treated AML-12 cells. To investigate the underlying mechanisms, GSTP1 overexpression, small interfering RNA, ferroptosis and Nrf2 inhibitors were used. GSTP1 methylation contributes to stress-induced hepatocellular damage and dysfunction. GSTP1 is involved in ferroptosis-mediated hepatocellular injury induced by restraint stress via the TF/Nrf2 pathway. These findings suggest that stress-induced hepatocellular injury is associated with ferroptosis, which is regulated by TF/Nrf2/GSTP1.

Peculiarities of the endocrine status of women with precancerous lesions of the vulva

The purpose of the study was to determine the hormonal profile of peripheral blood in women with precancerous lesions of the vulva.Material and methods. 309 women with precancerous lesions of the vulva and 60 gynecologically healthy women aged 25 to 70 years were included in the study. Patients with precancerous lesions were divided into 4 groups depending on the nosological unit: 87 women with severe vulvar dysplasia dependent on HPV (VHSIL), 154 individuals with differentiated vulvar dysplasia independent of HPV (dVIN), 36 patients with extramammary Paget's disease of the vulva (VPD) grade Ia and 32 women with melanoma in situ of the skin vulva In the groups with VHSIL, with dVIN, with melanoma in situ, there were women of premenopausal and postmenopausal age, therefore, for the comparison of hormonal data in these groups, subgroups were selected according to age: subgroups with persons younger than 50 years, subgroups of women aged 50 years and older. 60 conditionally healthy women of the control group also included 30 people under 50 years and 30 people 50 years and older. The levels of estradiol (E2), progesterone (P4), free testosterone (Tf), follicle-stimulating hormone (FSH), luteinizing hormone (LH), prolactin (PRL), anti-Müllerian hormone (AMH), insulin, thyroid-stimulating hormone (TSH), free triiodothyronine (T3f), free thyroxine (T4f), antibodies to thyroperoxidase (ATPO)in blood serumand HOMA indexwere determined. The results. There was no difference between serum LH and FSH levels in women with various precancerous diseases of the vulva under the age of 50 years. Individuals with dVIN were characterized by the highest level of PRL (24.44±1.55 ng/ml). Among women aged 50 years and older, the highest levels of LH, FSH and PRL were observed in persons with melanoma in situ - respectively 36.90±2.09 mIU/ml, 74.67±4.06 mIU/ml, 9.85±2.03 ng/ml; the level of FSH and PRL was higher, and the LH/FSH ratio was lower than in controls in all precancerous diseases of the vulva. No statistically significant difference was found between the basal levels of serum P4 and AMH in subgroups of women younger than 50 years and 50 years and older. Among both premenopausal and postmenopausal women, the highest average levels of E2 were recorded in women with melanoma in situ (104.61±10.22 pg/ml and 30.71±1.90 pg/ml), Tf and Tf index - in patients with dVIN (3.20±0.10 pg/ml and 1.58±0.05 pg/ml vs. 8.25±0.24% and 3.77±0.10%, respectively). Features of the thyroid status were the highest indicators of TSH and ATP in persons with dVIN both in premenopause (2.68±0.30 mIU/ml and 196.39±76.48 IU/ml) and in postmenopause (2.23±0 .09 mIU/ml and 123.46±16.07 IU/ml). The average level of 25(OH)D did not differ statistically significantly between women with various precancerous diseases of the vulva. The level of insulin and NOMA index among premenopausal women was the highest in women with melanoma in situ (14.33±1.55 mIU/ml), in postmenopausal women with VPD (14.90±1.02 mIU/ml), HOMA index – respectively, with dVIN (2.87±0.30) and with VPD (3.60±0.27). Insulin resistance was most common in premenopause among individuals with dVIN (53.33%), and in postmenopause - with VPD (72.22%).Conclusions. Features of the hormonal profile of people with precancerous diseases of the vulva under the age of 50 are the highest levels of PRL, TSH and ATPO in women with dVIN, the highest average levels of Tf and Tf index, insulin level and HOMA index – in patients with VPD, the highest average levels of E2– in women with melanoma in situ. Among postmenopausal patients, the highest levels of TSH and ATPO in women with dVIN, the highest average levels of Tf and Tf index, the level of insulin and NOMA index in patients with VPD, the highest average levels of LH, FSH, PRL and E2 in women with melanoma are characteristic features situ. The average level of 25(OH)D does not differ statistically significantly between women with various precancerous diseases of the vulva. Insulin resistance is most common in premenopausal individuals with dVIN, and in postmenopausal individuals with VPD.

Anti-Bacterial and Immunostimulatory Properties of Ulvan-Loaded Chitosan Nanoparticles for Use in Aquaculture.

Alternative prophylactic strategies to limit farm animal infection are needed in order to avoid the use of antibiotics. Anti-bacterial and immunostimulatory properties of bioactive compounds are of great interest in aquaculture. Marine derived polysaccharides, such as chitosan and ulvan, together with nanotechnology, have become the focus of attention in the scientific community due to their wide range of biological properties. In this work, chitosan and ulvan-loaded chitosan nanoparticles (referred as CS-TPP NPs and CS-UL-TPP NPs, respectively), obtained by the ionotropic gelation method, had round shape, and the mean sizes were 137.00 ± 5.44 and 325.50 ± 4.95nm, respectively. No study about the anti-bacterial activity of both types of NPs against Photobacterium damselae subsp. piscicida, an important fish pathogen, has been reported so far. Furthermore, the potential immunostimulatory effects of CS-UL-TPP NPs after oral administration in fish have not yet been evaluated. The percentage of bacterial inhibition against P. damselae subsp. piscicida was determined through in vitro assays, and it was significantly higher in CS-UL-TPP NPs than in CS-TPP NPs at concentrations below 0.03mgmL-1. The effects on the immune system of CS-TPP and CS-UL-TPP NPs were evaluated in Solea senegalensis juveniles at 30days after oral administration. Lysozyme activity as well as gene expression levels of il1b, il6, hamp1, tf and c3 was significantly higher in CS-UL-TPP NP-treated groups than in the controls, and no significant differences were observed in CS-TPP NP-treated groups. Thus, ulvan extracted from the macroalgae Ulva ohnoi could improve anti-bacterial and immunostimulant properties of CS-TPP NPs thereby making them suitable to be used as vaccine adjuvant or as immunostimulant.

Mitophagy contributes to zinc-induced ferroptosis in porcine testis cells.

Zinc (Zn) is a critical microelement for physiological process, but excess exposure can cause testicular dysfunction. However, the underlying mechanism of Zn-induced ferroptosis via regulating mitophagy is unknown. In this study, a total of 60 male weaned pigs were randomly divided into three groups and the content of Zn were 75mg/kg (control), 750mg/kg (Zn-I), 1500mg/kg (Zn-II). Meanwhile, testicular cells were treated with ZnSO4 (0, 50 and 100μM), and in combination of ZnSO4 (100μM) and ferrostation-1, ML-210, or 3-methyladenine for 24h. Our results verified that Zn could cause ferroptosis and lipid peroxidation, which were characterized by down-regulating level of SLC7A11, GPX4, and ferritin, and up-regulating levels of MDA, CD71, TF, and HMGB1 by Western blot, immunohistochemistry, immunofluorescence, peroxidase assay, et.ac. The opposite effect was shown after treatment with ferrostation-1 or ML-210. Meanwhile, the mitophagy-related proteins (PINK, Parkin, ATG5, LC3-II/LC3-I) were significantly upregulated in vivo and in vitro. Most importantly, 3-methyladenine observably relieved ferroptosis under Zn treatment through inhibiting mitophagy. Collectively, we demonstrated that mitophagy contributes to Zn-induced ferroptosis in porcine testis cells, providing a new insight into Zn toxicology.

Buyang Huanwu Decoction alleviates blood stasis, platelet activation, and inflammation and regulates the HMGB1/NF-κB pathway in rats with pulmonary fibrosis

Ethnopharmacological relevanceQi deficiency and blood stasis are identified to be pathological factors of pulmonary fibrosis (PF) in traditional Chinese medicine (TCM) theory. Buyang Huanwu Decoction (BYHWD) is a traditional Chinese prescription ameliorating Qi deficiency and blood stasis. Aim of the studyThe objective of this study was to investigate the anti-fibrosis effect of BYHWD and the potential molecular mechanism in rats. Materials and methodsBleomycin was used to construct PF rat models. 27 PF rats were randomly divided into three groups based on treatments: model group (saline solution, n = 9), low-dose BYHWD group (3.5 g/kg, n = 9), and high-dose BYHWD group (14.0 g/kg, n = 9). Moreover, 9 normal rats were used as the blank group. The blood viscosity, coagulation indexes (APTT, TT, PT, and FIB), platelet-related parameters (PLT, PDW, MPV, PCT, and PLCR), platelet microparticles (PMPs), and inflammatory factors (IL-2, IL-10, IL-1β, IL-6, IL-8, IL-17, IFN-γ, TNF-α, PAC-1, HMGB1, NF-κB, and TF) were determined. The lung tissue samples of rats were observed after hematoxylin-eosin (HE) staining. The full component analysis of the BYHWD extract was performed using the ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method. The signaling pathway included into the study was selected on the basis of bioinformatics analysis and the results of the phytochemical analysis. The expression levels of genes and proteins involved in the selected signaling pathway were detected. ResultsCompared to the blank group, the whole blood viscosity, PLR, PDW, MPV, PCT, PLCR, PMPs, and the levels of IL-1β, IL-6, IL-8, IL-17, TNF-α, PAC-1, HMGB1, NF-κB, and TF were increased, while the levels of IL-2 and IL-10 were decreased in the model group. Both low-dose BYHWD and high-dose BYHWD reversed these PF-induced effects in spite of the fact that low-dose BYHWD had no significant effect on the level of NF-κB. In addition, BYHWD ameliorated PF-induced inflammation in the rat lung tissue. The phytochemical analysis of the BYHWD extract combined with the bioinformatics analysis suggested that the therapeutical effect of BYHWD on PF was related to the HMGB1/NF-κB pathway, which consisted of NF-κB, IKBKB, ICAM1, VCAM1, HMGB1, and TLR4. Both RT-qPCR and western blot analyses showed that PF induced increases in the expression levels of NF-κB, ICAM1, VCAM1, HMGB1, and TLR4, but a decrease in the expression level of IKBKB. Moreover, both low-dose BYHWD and high-dose BYHWD exerted the opposite effects, and recovered the expression levels of NF-κB, ICAM1, VCAM1, HMGB1, TLR4, and IKBKB, despite the fact that low-dose BYHWD had no effects on the mRNA expression levels of NF-κB or TLR4. ConclusionsIn summary, BYHWD alleviated PF-induced blood stasis, platelet activation, and inflammation in the rats. Our study suggested BYHWD had a therapeutic effect on PF and was a good alternative for the complementary therapy of PF, and the potential molecular mechanism was modulation of HMGB1/NF-κB signaling pathway, and it needs further study.

TIPS-03 A PHASE 1 STUDY OF GALLIUM MALTOLATE (GAM) IN PATIENTS WITH RECURRENT GLIOBLASTOMA (GBM)

Abstract Glioblastoma remains the most aggressive malignant brain tumor with universally poor prognosis. New treatments are needed which target novelpathways. GBM cells have significantly greater requirements for iron and display markedly higher levels of transferrin receptors for iron uptake and ferritin for iron storage than surrounding normal astrocytes. The increased intracellular iron pool drives GBM activity. Previously, we showed that since gallium shares certain chemical properties with iron, it can inhibit malignant cell growth by disrupting tumor iron homeostasis. Recently, we reported that GaM is cytotoxic to GBM cell lines and stem cells but not to microvascular endothelial cells. In rodent orthotopically implanted human GBM xenografts, orally administered GaM reduced tumor volume and prolonged animal survival. These preclinical studies form the basis for our Phase 1 study which evaluates GaM in patients with recurrent GBM. The trial will follow a 3 + 3 phase 1 dose escalation design. Three to six patients at each dose level will receive GaM 500 mg/d, 1000 mg/d, or 1500 mg/day for a minimum of two 28-day cycles. Patients age >= 18 years with histologically confirmed GBM or molecular GBM with recurrent measurable disease are eligible. In the absence of measurable disease, pathologic confirmation of recurrence is required. Primary objectives are to determine a maximum-tolerated dose and recommended Phase 2 dose. Safety/tolerability will be assessed. Secondary objectives include disease assessments and evaluation of PFS and OS. Correlative studies planned are sequential blood levels of gallium, iron, Tf, ferritin, and hepcidin. Available tumor tissues will be examined for the expression iron biomarkers by immunochemistry. The study is currently enrolling. Patients will receive study drug until disease progression, unacceptable adverse events, patient withdrawal, or death. To date, 5 patients received GaM 500 mg/d without DLT and 4 patients started GaM 1000 mg/d.

Evaluation the Sensitivity and Specificity of Fucose and Some Biochemical Parameters as an Indicator of Therapeutic Response to Metformin in Polycystic Ovary Syndrome

The present study aimed to evaluate the sensitivity and specificity of fucose and some biochemical parameters, as an indicator of therapeutic response to Metformin in patients with recently diagnosed with Polycystic Ovary Syndrome-PCOS undergoing treatment. This study was conducted from 1/9/2022 to 1/11/2022. In 90 blood samples were collected from women aged between (35-18) years old. Of these, 30 samples were taken from women who had recently been diagnosed with polycystic ovary syndrome as a first group-G1, and another 30 samples were taken from the same group of women after taking metformin medication as second group-G2, compared to 30 samples taken from healthy women with no history of the disease (as a control group-C). Samples were collected from women's clinics and external laboratories (after diagnosis from the doctor) in Samarra city/ Salah Ul-Din. The study includes determination of the concentration of sex hormones(Follicle stimulating hormone-FSH, luteinizing hormone-LH, and testosterone) and some biochemical parameters(Galactose, blood glucose, total fucose-TF, and fucose binding protein-FBP) in sera of samples under investigation. There was a significant increase in the levels of FSH and Glucose in a G1 at a probability level (P≤0.05). However, there was a significant decrease in total fucose and FBP, with no-significant difference in the levels of LH, testosterone, Galactose as compared with the control group. There was a significant increase in G2 at a probability level (P≤0.05) in the levels of Testosterone, Galactose, glucose with a significant decrease in the levels of TF and FBP, with no-significant difference in the level of LH, FSH and transaminases enzymes as compared with the control group. The study also include study the Receiver operating characteristic-ROC , In G1 as compared with C showed that the Sensitivity was also recorded as highest for galactose , In G2 as compared with C showed that the area under curve was outstanding for galactose, Sensitivity was also recorded as highest for galactose , In G1 as compared with G2 showed that the area under curve was outstanding for galactose. Sensitivity was also recorded as highest for galactose, then for Testosterone and total fucose. From the above results we can conclude that the level of fucose may be used as a diagnostic biomarker for polycystic ovary syndrome and also for treatment response with metformin. We conclude from the current study that the level of fucose may be considered a diagnostic indicator for polycystic ovarian cysts, as well as an indicator of the therapeutic response to uremia in patients.

Upregulation of SGLT1 and 2 promotes oxidative stress in right atrial appendages of patients with low-grade inflammatory responses: potential role in atrial fibrillation

Abstract Introduction Atrial fibrillation (AF), the most common cardiac arrhythmia, is associated with increased morbidity and mortality. Inflammatory infiltrates have been observed in atrial tissues from AF patients suggesting that low-grade inflammation contributes to the pro-fibrotic, pro-remodeling and pro-senescence responses of the atrial tissue favoring AF. Although sodium glucose co-transporter2 inhibitors (SGLT2i) have shown beneficial effects in heart failure, the expression and role of SGLT2 in cardiac tissues remain unclear. Purpose This study examined the expression of SGLT1 and 2 in right atrial appendages (RAA) of cardiac patients, and, if so, to determine the underling mechanism and their functional role. Methods RAA were harvested from patients undergoing coronary artery bypass surgery or aortic valve replacement at the University Hospital of Strasbourg. Patients with comorbidities such as malignancies and chronic inflammatory diseases were excluded. The expression level of markers was assessed using RT-qPCR, Western blot analysis, and immunofluorescence staining, and the level of reactive oxygen species using dihydroethidium staining. Results Analysis of RAA samples evidenced up-to a 4-fold difference in the NOS3 mRNA level. Those with low levels of NOS3 mRNA showed high levels of ICAM1, F3, MMP9, TGF-β1, TP53, CDKN1A, CDKN2A, SLCA1, SLCA2, IL1B, IL6, TNF-a and CD68 whereas the contrary was observed in those with high levels of NOS3 mRNA. SGLT1/2 mRNA levels were negatively correlated with that of eNOS and positively with those of IL-1, IL-6, TNF- α, CD68, and the senescence marker p53. Western blot analysis of RAA indicated that those expressing low levels of eNOS protein showed high levels of ICAM-1, VCAM-1, TF, MMP-2, TGF-b, SGLT1, SGLT2, p-p65, p53, p21 and p16 proteins whereas the contrary was observed in those with high levels of eNOS protein. Immunofluorescence staining indicated that SGLT1/SGLT2 signals were colocalized with those of CD31, an endothelial cell marker, in RAA, and associated with those of CD68 and TNF-α. Samples with high levels of SGLT1 and 2 displayed higher levels of oxidative stress that were inhibited by the antioxidant NAC (N-acetylcysteine), VAS-2870 (NADPH oxidase inhibitor), perindoprilat (ACE inhibitor), losartan (AT1R antagonist), empagliflozin (selective SGLT2 inhibitor), sotagliflozin (dual SGLT1/2 inhibitor) and by infliximab (TNF-α receptor neutralizing antibody). Conclusion The findings indicate that high expression levels of SGLT1 and 2 are observed in RAA showing low eNOS levels associated with pro-inflammatory, pro-senescent, pro-remodeling and pro-fibrotic responses. They were further associated with increased levels of oxidative stress sensitive to inhibitors of either the local angiotensin system, TNF-alpha or SGLT2. Thus, inhibition of SGLT2 appears to be an interesting approach to reduce the stimulatory pro-oxidant signal in atrium related to low-grade inflammation. Funding Acknowledgement Type of funding sources: Private company. Main funding source(s): Boehringer Ingelheim Pharma GmbH & Co. KG

Clinical and bacterial outcomes of photodynamic therapy in the treatment of chronic necrotizing ulcerative periodontitis

AimTo assess the clinical periodontal and microbiological parameters in patients having chronic necrotizing ulcerative periodontitis (NUP) after the administration of adjunctive photodynamic therapy and non-surgical periodontal therapy in smokers, mal-nutrition and HIV positive individuals. Materials and MethodsA total of 30 individuals with NUP were selected for the present clinical trial, where both Group I and Group II had equal number of patients, respectively (15 each). The groups were divided on the basis of provision of treatment where patients in Group I underwent scaling and root planing (SRP). Furthermore, Group II patients were subjected to adjunctive phtotodynamic therapy and SRP (aPDT + SRP). Full mouth plaque scores (fmpS), full mouth bleeding on probing (fmBOP), periodontal pocket depth (PPD) and clinical attachment levels (CAL) were the clinical periodontal parameters that were carefully evaluated. Aggregatibacter actinomycetemcomitans (Aa), Porphyromonas gingivalis (Pg) and Tannerella forsythia (Tf) were the bacteria species which were evaluated. The assessments were done at baseline, three (3) months and six (6) months, respectively. ResultsAll periodontal parameters including fmpS, fmBOP, PPD and CAL significantly improved in both Group I and Group II, respectively. Group II patients subjected to aPDT + SRP reported higher reduction in mean PPD in comparison to Group I patients at follow up (p < 0.05). At follow-up, similar results were also reported for CAL gain where Group II (aPDT +SRP) patients reported higher CAL gain in comparison to patients who underwent SRP only (p < 0.05). From baseline to follow-up, all the bacterial levels exhibited reduction in both study groups i.e Group I (SRP) and Group II (aPDT + SRP), respectively (p < 0.05). However, Group II patients prominent reduction in the counts of Aa and Tf at the three-month interval, whereas Aa seem to reduce in HIV and smoking individuals at the six-month interval. Moreover, the levels of Pg and Tf significantly reduced at 3 months and only Aa at 6 months in patients with mal-nutrition, respectively (p < 0.05). ConclusionThe use of photodynamic treatment as an adjunct to scaling and root planing enhanced clinical periodontal results and reduced bacterial content in patients having NUP.

Abstract 6336: Regulation of tissue factor dependent procoagulant properties by CD44: Implication for metastasis of breast tumor cells

Abstract The aim here is to study the implication of the stem cell marker and fibrin receptor, CD44, in the regulation of tissue factor and its associated pro-coagulant and pro-metastatic properties in breast tumor cells. We examined this CD44/TF/coagulation axis in breast cancer EMT+ (MDA-MB-231, Hs578T), EMT- (MCF7, T47D) and EMT-inducible (EGF-treated MDA-MB-468, MDA-MB-468 harboring a doxycycline-inducible snail expression vector) cellular models. We used siRNA to inhibit CD44 expression and realized RT-qPCR, western blot, flow cytometry analyzes and promoter reporter assays to monitor regulations of our gene of interest. Furthermore, we realized in vitro clotting assays and TF activity assays to examine coagulant properties, and in vivo experimental metastasis assays to assess early metastatic potential. We first confirmed by WB, RT-qPCR and FACS analyzes that CD44 is over-expressed in our EMT+ and EMT-induced (EMTi) cellular models, concomitantly with TF expression. Using siRNAs, we showed that CD44 silencing in these same EMT+ and EMTi cellular models decreased both mRNA and protein levels of TF. Clotting assays and TF activity assays further showed that CD44 silencing decreased pro-coagulant properties of our EMT+ and EMTi cellular models in vitro and of their pro-metastatic properties in vivo. Examining further the molecular mechanisms of TF regulation by CD44, we employed truncated TF promoter reporter vectors to identify key transcription factors linking CD44 to TF transcription. These experiments revealed that a TF promoter region containing Sp1 binding sites is sufficient to mediate the diminution of TF promoter activity seen after CD44 silencing. Mutations in the Sp1 binding sites abrogated the activity of the TF reporter vector. Further pointing to Sp1 as a key transcription factor implicated in TF regulation by CD44, we observed that silencing CD44 inhibited Sp1 expression and that silencing Sp1 decreased TF mRNA levels. Our results thus demonstrate a key role of CD44 in regulating TF expression in EMT+ cancer cells, and point to Sp1 as a transcriptional mediator of this regulation. This modulation of TF expression by CD44 in EMT+ cells led to a concomitant regulation of pro-coagulant properties, and impacts metastatic potential. Citation Format: Amélie V. Villard, Anthony Genna, Christine Gilles. Regulation of tissue factor dependent procoagulant properties by CD44: Implication for metastasis of breast tumor cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6336.

Markers of activation of coagulation in cancer patients.

Prothrombotic tendency is characteristic of tumors. The aim of the study is to investigate the changes in the laboratory parameters for coagulation and fibrinolysis, namely in fibrinogen, thrombin-antithrombin complex (ТАТ), tissue factor (ТF), prothrombin fragment (F1+2), antithrombin III (AT III), D-dimer and screening coagulation tests in cancer patients before initiation of chemotherapy. Levels of F1+2, fibrinogen, ТАТ, AT III, TF, D-dimer, PT, aPTT and TT were measured baseline in 80 patients with breast and lung cancer before systemic treatment. The same parameters were investigated in 65 healthy volunteers. TF, ТАТ, F1+2 were measured by ELISA; AT III, D-dimer, fibrinogen and screening coagulation tests were measured by automated coagulation system Sysmex CS 2000i. RESULTS: Levels of F1+2, fibrinogen, ТАТ, TF, and D-dimer in cancer patients were significantly higher than those in the control group, while the levels of ATIII activity were significantly lower (p < 0.001). The highest area under the ROC curve was for D-dimer, which made it a good marker for the risk of thrombosis. Higher levels of TF, ТАТ, F1+2, fibrinogen and D-dimer and lower activity of АТ III in cancer patients support our hypothesis of an association between malignant disease and coagulation disorders. Cancer patients are at an increased risk of thrombosis wherefore antithrombotic prophylaxis may be considered (Tab. 6, Fig. 2, Ref. 34). Text in PDF www.elis.sk Keywords: coagulation, fibrinolysis, cancer.

The importance of association between sexsteroids deficiency, reduction of bone mineral density and falling risk in men with implications in medical rehabilitation

Introduction. Endocrino-metabolic rehabilitation represent one of the most complex sector in clinical medicine, regarding functional rehabilitation. Sex hormones deficiency plays an important role in the etiology of osteoporosis in men. At the same time, with age, the trophic role of androgens on muscle decreases and determines an increased frequency of falls. The objective of our study is to determine the association between sexsteroids deficiency, reduction of bone mineral density (BMD) and falling risk in men. Methods. Our retrospective cross-sectional study included 146 men aged between 65–85 years with low BMD (study group) and 121 men with normal BMD (control group). The measurement of Total testosterone (Tt), free testosterone (Tf) and estradiol (E2) serum levels was performed using the immunoassay or the immunoenzymatic methods. Femoral neck and lumbar spine BMD was determined using Dual-energy X-ray absorptiometry (DEXA). The risk of falls was assessed by Tandem Standing, Up &amp; Go, Chair – Rising and walking speed tests. Results. We found a significantly association between Tf and E2 deficiency and low BMD (p=0.007). Also, in men with reduced BMD (study group) we observed significant lower levels of Tf (p&lt;0.001) and E2 (p=0.003) compared to control group. E2 deficiency was associated significantly with low BMD and increased fall risk (p=0.001). At the same time the results highlighted significant lower levels of Tf in patients with BMD reduction and increased risk of falls (p=0.002). Tt deficiency was not associated with BMD reduction (p=0.088) or increased risk of falling (p=0.277). Conclusions. This research revealed a significant association between male sexsteroids deficiency, low BMD and increase of falling risk, with implications in rehabilitation program. The risk of ostoporosis and for falling in man can be estimated by determining serum Tf and E2 levels. Keywords: sexsteroids deficiency; bone mineral density; falling risk,

Estradiol modulated differentiation and dynamic growth of CD90+ spermatogonial stem cells toward Sertoli-like cells

Mouse CD90+ SSCs were enriched using the MACS technique and incubated with different doses of estradiol, ranging from 0.01 ng/mL to 500 μg/mL, for 7 days. The viability of SSCs was determined using an MTT assay. The combined effects of estradiol plus Sertoli cell differentiation medium on the orientation of SSCs toward Sertoli-like cells were also assessed. Using immunofluorescence imaging, we monitored protein levels of Oct3/4 after being exposed to estradiol. In addition, protein levels of testosterone, TF, and ABP were measured using ELISA. The expression of Sertoli cell-specific genes such as SOX9, GATA4, FSHR, TF, and ESR-1 and -2 was monitored using real-time PCR assay, and the effects of 14-day injection of estradiol on sperm parameters and Oct3/4 positive progenitor cells in a model of mouse were determined.Data showed that estradiol increased the viability of mouse SSCs in a dose-dependent manner compared to the control (p < 0.05). Along with these changes, cells displayed morphological changes and reduced Oct3/4 transcription factor levels compared to the control SSCs. 7-day incubation of SSCs with estradiol led to the up-regulation of SOX9, GATA4, FSHR, TF, and ESR-1 and -2, and levels of testosterone, TF, and ABP were increased compared to the control group (p < 0.05). The in-vivo examination noted that estradiol reduced sperm parameters coincided with morphological abnormalities (p < 0.05). Histological examination revealed pathological changes in seminiferous tubules and reduction of testicular Oct3/4+ progenitor cells.In conclusion, estradiol treatment probably can induce Sertoli cell differentiation of SSCs while exogenous administration leads to testicular progenitor cell depletion and infertility in long term.

Different Profiles of Cytokines, Chemokines and Coagulation Mediators Associated with Severity in Brazilian Patients Infected with Dengue Virus.

The incidence of dengue in Latin America has increased dramatically during the last decade. Understanding the pathogenic mechanisms in dengue is crucial for the identification of biomarkers for the triage of patients. We aimed to characterize the profile of cytokines (IFN-γ, TNF-α, IL-1β, IL-6, IL-18 and IL-10), chemokines (CXCL8/IL-8, CCL2/MCP-1 and CXCL10/IP-10) and coagulation mediators (Fibrinogen, D-dimer, Tissue factor-TF, Tissue factor pathway inhibitor-TFPI and Thrombomodulin) during the dengue-4 epidemic in Brazil. Laboratory-confirmed dengue cases had higher levels of TNF-α (p < 0.001), IL-6 (p = 0.005), IL-10 (p < 0.001), IL-18 (p = 0.001), CXCL8/IL-8 (p < 0.001), CCL2/MCP-1 (p < 0.001), CXCL10/IP-10 (p = 0.001), fibrinogen (p = 0.037), D-dimer (p = 0.01) and TFPI (p = 0.042) and lower levels of TF (p = 0.042) compared to healthy controls. A principal component analysis (PCA) distinguished between two profiles of mediators of inflammation and coagulation: protective (TNF-α, IL-1β and CXCL8/IL-8) and pathological (IL-6, TF and TFPI). Lastly, multivariate logistic regression analysis identified high aspartate aminotransferase-to-platelet ratio index (APRI) as independent risk factors associated with severity (adjusted OR: 1.33; 95% CI 1.03–1.71; p = 0.027), the area under the receiver operating characteristics curve (AUC) was 0.775 (95% CI 0.681–0.869) and an optimal cutoff value was 1.4 (sensitivity: 76%; specificity: 79%), so it could be a useful marker for the triage of patients attending primary care centers.

The Performance of Immunoassays to Measure Antibodies to the Chlamydia trachomatis Antigen Pgp3 in Different Epidemiological Settings for Trachoma.

ABSTRACT.Programs to eliminate trachoma as a public health problem use prevalence of the clinical sign trachomatous inflammation—follicular (TF) in 1- to 9-year-olds in endemic districts to make decisions to begin or end mass drug administration with azithromycin. Trachomatous inflammation—follicular is used as a proxy for transmission of ocular Chlamydia trachomatis infection. Long-term monitoring of previously endemic districts for recrudescence of ocular C. trachomatis infection would benefit from a simple blood test that could be integrated with other public health programs. In this study, we evaluated multiple tests to measure antibodies against the C. trachomatis antigen Pgp3—a multiplex bead assay (MBA), an ELISA, and two versions of a lateral flow assay (LFA)—in four districts of the Amhara region of Ethiopia with varying levels of TF. Seroprevalence and seroconversion rate (SCR) results were proportional to TF prevalence by district for most tests, with the notable exception of the LFA using colloidal gold as the developing reagent. Changing the test developing reagent to black latex improved agreement between serological measures and TF prevalence and in inter-rater agreement. Seroconversion rate estimates using data derived from the LFA-gold assay were inconsistent with the shape of the age-seroprevalence curve, which did not increase in older ages. These data revealed potential complications with using SCR that will need further evaluation. Data from MBA, ELISA, and LFA with the black test line showed good agreement with each other and proportionality to TF estimates, providing further data that serology has potential utility for trachoma surveillance.

Tissue Factor Expression As a Prognostic Marker in Patients with B-Cell Malignant Lymphoma

Background. Studies in tumor cell lines have shown an increased expression of TF. Expression of TF in adenocarcinoma of the pancreas is associated with tumor progression. It has been described that between 5% and 10% of patients with malignant lymphoma (ML) may develop VTD. The mechanisms of ML-associated thrombosis are unknown, however, expression of functional TF may increase as much as 75-fold in peripheral blood monocytes, particularly during relapse. As expected, thrombosis was found in patients with the highest TF levels. The role of TF in the biology of ML has not yet been determined. Considering indirect evidence, it seems that, besides its procoagulant effects, TF may have a role in tumor progression.Objective. The purpose of the present study was to elucidate whether the interaction of tissue factor and TF/VIIa could induce relapsed in non Hodgkin Lymphoma patients.Patients and Methods. We therefore performed a prospective study (cohort) to evaluate the risk of TF in non Hodgkin Lymphoma patients without a previous episode of venous thromboembolism. 59 patients 30 women and 29 men were enrolled in this study. The median age was 43.3 ± 14.5 (range 20 to 59). The TF and FT/VIIa activity was performed on fresh plasma (American Diagnostics, USA), and other tests (PC, PS, AT and activated protein C resistance) (STAGO, Asnieres, Francia).Results.59 patients with B-cell ML, 30 women and 29 men with a mean age of 43.3 years (ranges 20 to 59). According to the Ann Arbor Staging system, 44% percent of patients (n=26) had a stage IV and only 14% at stage I. ML was mostly limited to lymph nodes (74%).Results of hemostatic markers were between the normal ranges. However, concentrations of factors involved in the initiation phase of fluid phase of blood coagulation (TF, FT/VIIa complex, and FVII) had an abnormal distribution. None case with APCR was found.Comparison between complete remission and relapsing patients. Thirty-one patients (52.5%) achieved complete remission at the end of follow-up and 28 (47.5%) relapsed. Mean follow up for patients with complete remission was 48 months vs. 18 months for relapsing patients (p <0.001). After comparing TF concentrations between the groups, relapsing patients had higher levels of TF (p=0.005). TF/FVIIa complex levels were also significantly higher in patients with relapsing disease (p=0.019). As expected LDH levels were significantly lower in patients with complete remission (p=0.0001).TF levels and response to treatment. Twenty-nine patients had normal TF levels. Relapses were more frequently found in patients with elevated TF concentrations (n=19) as compared with patients with normal TF levels (n=9) (p=0.013). Elevated TF/FVIIa complexes levels were also more frequently seen among patients with relapsing ML (n=32) as compared with patients with complete remission (n=27) (p=0.046).As expected, other factors associated with a poor prognosis were DHL levels (p=0.001) and IPI (p=0.001).Discussion. TF and TF/FVIIIa concentrations were significantly different in patients who achieved complete remission as compared with those with relapse after treatment. As expected for the staging of patients included LDH serum levels as well as IPI score were also significantly different between these two groups of patients. Moreover, estimation of the relative risks associated with relapsing disease showed that also these four variables were associated with relapsing disease. However, linear regression analysis showed that TF, LDH, and IPI but not TF/FVIIa complexes remained as a risk factor for relapse in patients with ML. To our knowledge, we have demonstrated for the first time that high TF concentration is a prognosis factor for relapse in patients with B-cell ML.Our results seem consistent with previous reports regarding an association between the increased TF expression and advanced staging of the malignant disease. It has been demonstrated the presence of high plasma levels of TF and FVIII in patients with solid tumors suggesting that it may be an important mechanism to activate blood coagulation system in cancer patients. Tumor cells may also induce TF expression on the cell surface of peripheral blood monocytes and macrophages. DisclosuresNo relevant conflicts of interest to declare.