Hasil untuk "Cytology"

Sharif Alhajlah, Adam M Thompson, Zubair Ahmed

In the original manuscript [...]

Beatriz Araújo, Inês Serrenho, Andreia Valente da Silva et al.

Mesenchymal stem cells (MSCs) exhibit unique properties that make them promising candidates for cell therapy, particularly in neurological disorders. They can be derived from various tissues, with bone marrow, adipose tissue, umbilical cord, and placenta being the most common sources. Evidence suggests that the tissue of origin significantly influences MSC characteristics, including secretome composition, proliferation rate, and adhesion capacity.Clinical trials have demonstrated the safety and therapeutic potential of MSCs in conditions such as spinal cord injury, multiple sclerosis, and stroke. MSC therapy has been associated with improvements in motor, sensory, and cognitive functions, as well as enhanced quality of life. Mechanistically, MSCs promote neuroprotection, reduce inflammation, and modulate immune responses. In spinal cord injury, intrathecal administration of adipose- and bone marrow-derived MSCs has led to significant functional recovery, with single high-dose treatments often yielding better outcomes than multiple lower doses. In amyotrophic lateral sclerosis, bone marrow-derived MSCs have shown potential in slowing disease progression, though higher doses do not always result in greater benefits. In multiple sclerosis, high doses of umbilical cord-derived MSCs improved quality of life and prevented disease progression, whereas lower doses of bone marrow-derived MSCs provided limited functional benefits.While MSC therapy is considered safe, patient responses vary, and a definitive correlation between administered dose and therapeutic effects remains elusive. The small number of studies using comparable protocols impedes comparison of other relevant factor, limits the drawing of conclusions and underscore the importance of developing standardized protocols to optimize MSC-based treatments and maximize their clinical efficacy.

Esther García-Rojo, Javier Angulo, Mariam El Assar et al.

We aimed to determine the influence of donors’ vascular function on renal function in recipients and to evaluate the role of Orai1 calcium channels as a potential marker. A prospective collaborative multicenter study was designed. Blood, aorta (HA), mesenteric arteries (HMAs) and corpus cavernosum (HCC) specimens were obtained from organ donors at the kidney procurement procedure (n = 60). Evolution (up to 2 years) of renal function measured as serum creatinine (SCr) and glomerular filtration rate (GFR) was evaluated in respective recipients (n = 64). Vascular responses were determined in HA, HMA and HCC from donors. Tumor necrosis factor-α, asymmetric dimethyl arginine and Orai1 were measured in plasma. Orai1 protein expression was also evaluated in each donor’s aorta. Endothelium-dependent vasodilation (HMA, HCC) and adrenergic contraction (HA) in donors determined renal function in recipients, 12 months post-transplantation. Donors in the best quartile of vascular function predicted lower SCr and higher GFR in kidney recipients for 12/24 months. Plasma Orai1 in donors was negatively correlated with vascular function and predicted renal function at 3–6 months post-transplantation. Donor Orai1 vascular content was associated with reduced vascular function and with poorer recipient renal function for 1-year post-transplantation. Systemic vascular function of kidney donors determines recipients’ renal function at short/mid-term. Donors’ vascular function and recipients’ renal function are negatively associated with donors’ Orai1 vascular expression, being a potential biomarker of renal outcomes.

Lakshana Sruthi Sadu Murari, Sam Kunkel, Anala Shetty et al.

The p63 protein is a master regulatory transcription factor that plays crucial roles in cell differentiation, adult tissue homeostasis, and chromatin remodeling, and its dysregulation is associated with genetic disorders, physiological and premature aging, and cancer. The effects of p63 are carried out by two main isoforms that regulate cell proliferation and senescence. p63 also controls the epigenome by regulating interactions with histone modulators, such as the histone acetyltransferase p300, deacetylase HDAC1/2, and DNA methyltransferases. miRNA-p63 interactions are also critical regulators in the context of cancer metastasis. This review aims to elaborate on the diverse roles of p63, focusing on disease, development, and the mechanisms controlling genome organization and function.

Yaping Qin, Mingqing Yang, Yunzhu Cao et al.

Abstract Nasopharyngeal carcinoma (NPC) is a highly aggressive head and neck cancer characterized by a complex etiology and a propensity for metastasis. The current study explores the intricate relationship between Forkhead Box A1 (FOXA1) and B-cell-specific Moloney murine leukemia virus integration site 1 (BMI1) in the cancer progression and chemoresistance of NPC. Our research identified a significant downregulation of FOXA1 in NPC tissues and cell lines, which correlates with advanced clinical stages and poor differentiation, underscoring its potential role as a tumor suppressor. Functional assays demonstrated that the silencing of FOXA1 significantly enhanced the proliferation, migration, and invasive capabilities of NPC cells in vitro. Furthermore, the deficiency of FOXA1 was associated with a diminished sensitivity to cisplatin, as evidenced by increased cell viability, reduced apoptosis, and impaired cell cycle arrest upon drug exposure. Mechanistic studies revealed BMI1 as a critical downstream target of FOXA1. We observed a negative correlation between the expression levels of FOXA1 and BMI1 in NPC tissues. FOXA1 was shown to bind directly to the BMI1 promoter, effectively dampening its transcriptional activity. Rescue experiments indicated that the downregulation of BMI1 could partially reverse the malignant phenotypes induced by FOXA1 silencing, both in vitro and in vivo. Importantly, the knockdown of BMI1 significantly increased the chemosensitivity of FOXA1-depleted NPC cells to cisplatin, effectively counteracting the drug resistance associated with FOXA1 suppression. These findings highlight the pivotal role of FOXA1 in NPC development and progression and suggest that its loss leads to the upregulation of BMI1 and the acquisition of cisplatin resistance. Our study provides novel insights into the molecular mechanisms underlying the malignancy and chemoresistance of NPC and proposes that targeting the FOXA1/BMI1 axis could offer a promising therapeutic strategy for the treatment of this devastating disease.

Jacinda Tran, Christine Lee Hathaway, Cara Jill Broshkevitch et al.

IntroductionWomen living with human immunodeficiency virus (WLHIV) face elevated risks of human papillomavirus (HPV) acquisition and cervical cancer (CC). Coverage of CC screening and treatment remains low in low-and-middle-income settings, reflecting resource challenges and loss to follow-up with current strategies. We estimated the health and economic impact of alternative scalable CC screening strategies in KwaZulu-Natal, South Africa, a region with high burden of CC and HIV.MethodsWe parameterized a dynamic compartmental model of HPV and HIV transmission and CC natural history to KwaZulu-Natal. Over 100 years, we simulated the status quo of a multi-visit screening and treatment strategy with cytology and colposcopy triage (South African standard of care) and six single-visit comparator scenarios with varying: 1) screening strategy (HPV DNA testing alone, with genotyping, or with automated visual evaluation triage, a new high-performance technology), 2) screening frequency (once-per-lifetime for all women, or repeated every 5 years for WLHIV and twice for women without HIV), and 3) loss to follow-up for treatment. Using the Ministry of Health perspective, we estimated costs associated with HPV vaccination, screening, and pre-cancer, CC, and HIV treatment. We quantified CC cases, deaths, and disability-adjusted life-years (DALYs) averted for each scenario. We discounted costs (2022 US dollars) and outcomes at 3% annually and calculated incremental cost-effectiveness ratios (ICERs).ResultsWe projected 69,294 new CC cases and 43,950 CC-related deaths in the status quo scenario. HPV DNA testing achieved the greatest improvement in health outcomes, averting 9.4% of cases and 9.0% of deaths with one-time screening and 37.1% and 35.1%, respectively, with repeat screening. Compared to the cost of the status quo ($12.79 billion), repeat screening using HPV DNA genotyping had the greatest increase in costs. Repeat screening with HPV DNA testing was the most effective strategy below the willingness to pay threshold (ICER: $3,194/DALY averted). One-time screening with HPV DNA testing was also an efficient strategy (ICER: $1,398/DALY averted).ConclusionsRepeat single-visit screening with HPV DNA testing was the optimal strategy simulated. Single-visit strategies with increased frequency for WLHIV may be cost-effective in KwaZulu-Natal and similar settings with high HIV and HPV prevalence.

Tamer Mohamed Abo-Sriea, Elshaimaa Ismael, Basant Mohsen Sobhi et al.

Stress in poultry production is energy-demanding. Nucleotides and yeast cell-wall products are essential nutrients for broiler performance, gut function, and immune response. Antibiotics, like florfenicol, negatively affect the immune system. A total of 600 one-d-old broiler chickens (Cobb-500) were weighed and randomly allotted into four groups with three replicates each. The control group (G1) received the basal diet, G2 received a diet supplemented with a combination of nucleotides and Saccharomyces cerevisiae derivatives (250 g/Ton), G3 received the basal diet and medicated with florfenicol (25 mg/Kg body weight) in drinking water for 5 days, while G4 received a combination of nucleotides and Saccharomyces cerevisiae-derivatives (250 g/Ton) and medicated with florfenicol in drinking water. Growth performance criteria were recorded weekly. Blood, intestinal contents, small-intestine sections, and litter samples were collected to measure birds’ performance, carcass yields, leukocytic counts, antioxidant capacity, antibody titres, phagocytic index, caecal Clostridia, intestinal histomorphometry, and litter hygiene. Nucleotide-supplemented groups (G2 and G4) revealed significant (p ≤ 0.05) improvements in feed conversion, and body weight, but not for carcass yields in comparison to the control. Dietary nucleotides in G2 elevated blood total proteins, leucocytic count, antioxidant capacity, and phagocytic index, while they lowered blood lipids and litter moisture and nitrogen (p ≤ 0.05). Dietary nucleotides in G4 ameliorated the immunosuppressive effect of florfenicol (p ≤ 0.05) indicated in reducing caecal Clostridia, improving duodenal and ileal villi length, and increasing blood albumin and globulin levels, and phagocytosis%. Supplementing diets with nucleotides and yeast products has improved the immune system and provided a healthier gut for broilers.

Andrea Pagani, Dominik Duscher, Sebastian Geis et al.

Background: Extracellular vesicles are pivotal mediators in intercellular communication, facilitating the exchange of biological information among healthy, pathological and tumor cells. Between the diverse subtypes of extracellular vesicles, exosomes have unique properties and clinical and therapeutical applications. Breast cancer ranks as one of the most prevalent malignancies across the globe. Both the tumor core and its surrounding microenvironment engage in a complex, orchestrated interaction that facilitates cancer’s growth and spread. Methods: The most significant PubMed literature about extracellular vesicles and Adipose-Derived Stem Cell Exosomes and breast cancer was selected in order to report their biological properties and potential applications, in particular in treating triple-negative breast cancer. Results: Adipose-Derived Stem Cell Exosomes represent a potential tool in targeting triple-negative breast cancer cells at three main levels: the tumor core, the tumor microenvironment and surrounding tissues, including metastases. Conclusions: The possibility of impacting triple-negative breast cancer cells with engineered Adipose-Derived Stem Cell Exosomes is real. The opportunity to translate our current in vitro analyses into a future in vivo scenario is even more challenging.

Chan Ho Noh, Sangryong Park, Hye-Rim Seong et al.

Tissue regeneration is an essential requirement for wound healing and recovery of organs’ function. It has been demonstrated that wound healing can be facilitated by activating paracrine signaling mediated by exosomes secreted from stem cells, since exosomes deliver many functional molecules including growth factors (GFs) and neurotrophic factors (NFs) effective for tissue regeneration. In this study, an exosome-rich conditioned medium (ERCM) was collected from human amniotic membrane stem cells (AMSCs) by cultivating the cells under a low oxygen tension (2% O₂ and 5% CO₂). The contents of GFs and NFs including keratinocyte growth factor, epidermal growth factor, fibroblast growth factor 1, transforming growth factor–β, and vascular endothelial growth factor responsible for skin regeneration were much higher (10–30 folds) in the ERCM than in normal conditioned medium (NCM). In was found that CM–DiI-labeled exosomes readily entered keratinocytes and fibroblasts, and that ERCM not only facilitated the proliferation of keratinocytes in normal condition, but also protected against H₂O₂ cytotoxicity. In cell-migration assay, the scratch wound in keratinocyte culture dish was rapidly closed by treatment with ERCM. Such wound-healing effects of ERCM were confirmed in a rat whole skin-excision model: i.e., the wound closure was significantly accelerated, remaining minimal crusts, by topical application of ERCM solution (4 × 10⁹ exosome particles/100 μL) at 4-day intervals. In the wounded skin, the deposition of collagens was enhanced by treatment with ERCM, which was supported by the increased production of collagen-1 and collagen-3. In addition, enhanced angiogenesis in ERCM-treated wounds was confirmed by increased von Willebrand factor (vWF)-positive endothelial cells. The results indicate that ERCM from AMSCs with high concentrations of GFs and NFs improves wound healing through tissue regeneration not only by facilitating keratinocyte proliferation for skin repair, but also activating fibroblasts for extracellular matrix production, in addition to the regulation of angiogenesis and scar tissue formation.

Kara Chandler, Josh Millar, George Ward et al.

Extracellular vesicles (EVs) are lipid bilayer structures released by all cells that mediate cell-to-cell communication via the transfer of bioactive cargo. Because of the natural origin of EVs, their efficient uptake by recipient cells, capacity to stabilize and transport biomolecules and their potential for cell/tissue targeting and preferential uptake by cancer cells, they have enormous potential for bioengineering into improved and targeted drug delivery systems. In this work, we investigated the use of laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) as a tool to measure the loading of platinum-based chemotherapeutic agents. The EV loading of oxaliplatin via co-incubation was demonstrated, and LA-ICP-MS imaging showed greater efficiency of delivery to colorectal cancer cells compared to free oxaliplatin, leading to enhanced cytotoxic effect. Further, the impact of EV co-loading with a porphyrin (C5SHU, known as ‘C5’) photosensitizer on oxaliplatin delivery was assessed. Fluorescence analysis using nano-flow cytometry showed dose-dependent EV loading as well as a trend towards the loading of larger particles. Exposure of OXA-C5-EV-treated colorectal cancer cells to light indicated that delivery was enhanced by both light exposure and porphyrins, with a synergistic effect on cell viability observed between oxaliplatin, EVs and light exposure after the delivery of the co-loaded EVs. In summary, this work demonstrates the utility of LA-ICP-MS and mass spectrometry imaging in assessing the loading efficiency and cellular delivery of platinum-based therapeutics, which would also be suitable for agents containing other elements, confirms that EVs are more efficient at delivery compared to free drugs, and describes the use of light exposure in optimizing delivery and therapeutic effects of EV-mediated drug delivery both in combination and independently of porphyrin-based photosensitizers.

Ya’nan Qu, Rifeng Gao, Xiang Wei et al.

Abstract Cardiomyocyte pyroptosis and apoptosis play a vital role in the pathophysiology of several cardiovascular diseases. Our recent study revealed that gasdermin D (GSDMD) can promote myocardial I/R injury via the caspase-11/GSDMD pathway. We also found that GSDMD deletion attenuated myocardial I/R and MI injury by reducing cardiomyocyte apoptosis and pyroptosis. However, how GSDMD mediates cardiomyocyte apoptosis and protects myocardial function remains unclear. Here, we found that doxorubicin (DOX) treatment resulted in increased apoptosis and pyroptosis in cardiomyocytes and that caspase-11/GSDMD could mediate DOX-induced cardiotoxicity (DIC) injury. Interestingly, GSDMD overexpression promoted cardiomyocyte apoptosis, which was attenuated by GSDMD knockdown. Notably, GSDMD overexpression exacerbated DIC injury, impaired cardiac function in vitro and in vivo, and enhanced DOX-induced cardiomyocyte autophagy. Mechanistically, GSDMD regulated the activity of FAM134B, an endoplasmic reticulum autophagy receptor, by pore formation on the endoplasmic reticulum membrane via its N-terminus, thus activating endoplasmic reticulum stress. In turn, FAM134B interacted with autophagic protein LC3, thus inducing cardiac autophagy, promoting cardiomyocyte apoptosis, and aggravating DIC. These results suggest that GSDMD promotes autophagy and induces cardiomyocyte apoptosis by modulating the reaction of FAM134B and LC3, thereby promoting DIC injury. Targeted regulation of GSDMD may be a new target for the prevention and treatment of DIC.

Gustavo J. C. Freitas, Ludmila Gouveia-Eufrasio, Eluzia C. P. Emidio et al.

The phenotypic plasticity of <i>Cryptococcus neoformans</i> is widely studied and demonstrated in vitro, but its influence on pathogenicity remains unclear. In this study, we investigated the dynamics of cryptococcal cell and transcriptional remodeling during pulmonary infection in a murine model. We showed that in <i>Cryptococcus neoformans</i>, cell size reduction (cell body ≤ 3 µm) is important for initial adaptation during infection. This change was associated with reproductive fitness and tissue invasion. Subsequently, the fungus develops mechanisms aimed at resistance to the host’s immune response, which is determinant for virulence. We investigated the transcriptional changes involved in this cellular remodeling and found an upregulation of transcripts related to ribosome biogenesis at the beginning (6 h) of infection and a later (10 days) upregulation of transcripts involved in the inositol pathway, energy production, and the proteasome. Consistent with a role for the proteasome, we found that its inhibition delayed cell remodeling during infection with the H99 strain. Altogether, these results further our understanding of the infection biology of <i>C. neoformans</i> and provide perspectives to support therapeutic and diagnostic targets for cryptococcosis.

Jia Wang, Yue Sun, Yiping Liu et al.

Abstract Background The existence of mesenchymal stem cells (MSCs) in Schneiderian membrane has not been determined. The aim of this study is to investigate whether there are MSCs in Schneiderian membrane, and the effect of platelet-rich fibrin (PRF) on osteogenic differentiation of these cells and on new bone formation in maxillary sinus after maxillary sinus floor elevation. Methods Schneiderian membrane derived mesenchymal stem cells (SM-MSCs) were isolated from rabbit maxillary sinus. Cells were identified by flow cytometry and multipotential differentiation. Real-time cell analysis assay, fluorescence staining, transwell assay, and wound healing assay were used to determine the effects of PRF stimulation on cell proliferation and migration. The osteogenic differentiation ability of cells stimulated by PRF or osteoinductive medium was evaluated by alkaline phosphatase staining, alizarin red staining, PCR and Western blot. Equivalent volume Bio-oss and the mixture of Bio-oss and PRF were used as bone graft materials for maxillary sinus floor elevation. Micro-CT, bone double-staining, HE staining, Masson staining, and toluidine blue staining were used to evaluate the osteogenic effect in 8 and 12 weeks after surgery. Results The cell surface markers were positive for expression of CD90, CD105, and negative for expression of CD34, CD45. SM-MSCs had the ability of osteogenic, adipogenic and chondrogenic differentiation. PRF could stimulate proliferation, migration and osteogenic differentiation of SM-MSCs, which was achieved by up-regulating ERK 1/2 signaling pathway. PRF could accelerate the formation of new bone in maxillary sinus and increase the amount of new bone formation. Conclusions MSCs existed in Schneiderian membrane, and PRF stimulation could promote cell proliferation, migration and osteogenic differentiation. The application of PRF in maxillary sinus floor elevation could accelerate bone healing and increase the quantity and quality of new bone. PRF, as autologous graft materials, might offer a promising strategy for the clinical bone formation during MSFE procedure. Graphical abstract Video Abstract.

Syed Sarim Imam, Fahad A. Al-Abbasi, Salman Hosawi et al.

Platelet-rich plasma (PRP) is a widely accepted treatment approach and has heightened the quality of care among physicians. PRP has been used over the last decade to boost clinical results of plastic therapies, periodontal surgery and intra-bony defects. According to certain research, elevated levels of PRP growth factors that could promote tissue repair and have the potential for PRP to be beneficial in regenerating processes that Maxillofacial and Oral Surgeons, Veterinary Officers, Athletic medicine specialists and Dermatologists have long admired. PRP is an autologous whole blood fraction that has a heavy amount of a variety of growth factors such as epidermal growth factor (EGF), Vascular Endothelial Growth Factor (VEGF), hepatocyte growth factor (HGF), fibroblast growth factors (FGFs), transforming growth factor beta-1 (TGF-b), insulin-like growth factor-I (IGF-I) and platelet-derived growth factor (PDGF) which can facilitate repair and regeneration. Moreover, a clinical trial of PRP in severe angina patients has shown its excellent safety profile. However, PRP is a very complex biological substance with an array of active biomolecules, its functions are yet to be fully clarified. In-addition, there was insufficient work assessing possible cardiovascular tissue benefits from PRP. Thus, it still remains necessary to identify the most clinically important cardiovascular applications and further research in clinical scenario need to be validated.

Diana Sousa, Raquel T. Lima, Vanessa Lopes-Rodrigues et al.

Cancer multidrug resistance (MDR) is one of the main challenges for cancer treatment efficacy. MDR is a phenomenon by which tumor cells become resistant to several unrelated drugs. Some studies have previously described the important role of extracellular vesicles (EVs) in the dissemination of a MDR phenotype. EVs’ cargo may include different players of MDR, such as microRNAS and drug-efflux pumps, which may be transferred from donor MDR cells to recipient drug-sensitive counterparts. The present work aimed to: (i) compare the ability of drug-sensitive and their MDR counterpart cells to release and capture EVs and (ii) study and relate those differences with possible distinct fate of the endocytic pathway in these counterpart cells. Our results showed that MDR cells released more EVs than their drug-sensitive counterparts and also that the drug-sensitive cells captured more EVs than their MDR counterparts. This difference in the release and capture of EVs may be associated with differences in the endocytic pathway between drug-sensitive and MDR cells. Importantly, manipulation of the recycling pathway influenced the response of drug-sensitive cells to doxorubicin treatment.

Yang Li, Shuting Yang, Ling Qin et al.

Abstract Chondrogenesis is a major contributor to skeletal development and maintenance, as well as bone repair. Transcriptional coactivator with PDZ-binding motif (TAZ) is a key regulator of osteogenesis and adipogenesis, but how TAZ regulates chondrogenesis and skeletal development remains undefined. Here, we found that TAZ expression is gradually increased during chondrogenic differentiation. Deletion of TAZ in chondrocyte lineage impaired articular and growth plate, as well as the bone development in TAZ-deficient mice. Consistently, loss of TAZ impaired fracture healing. Mechanistically, we found that ectopic expression of TAZ markedly promoted chondroprogenitor proliferation, while deletion of TAZ impaired chondrocyte proliferation and differentiation. TAZ associated with Sox5 to regulate the expression and stability of Sox5 and downstream chondrocyte marker genes’ expression. In addition, overexpression of TAZ enhanced Col10a1 expression and promoted chondrocyte maturation, which was blocked by deletion of TAZ. Overall, our findings demonstrated that TAZ is required for skeletal development and joint maintenance that provided new insights into therapeutic strategies for fracture healing, heterotopic ossification, osteoarthritis, and other bone diseases.

Makoto Kato, Shin Tsunekawa, Nobuhisa Nakamura et al.

Diabetes is a major risk factor for atherosclerosis and ischemic vascular diseases. Recently, regenerative medicine is expected to be a novel therapy for ischemic diseases. Our previous studies have reported that transplantation of stem cells promoted therapeutic angiogenesis for diabetic neuropathy and ischemic vascular disease in a paracrine manner, but the precise mechanism is unclear. Therefore, we examined whether secreted factors from stem cells had direct beneficial effects on endothelial cells to promote angiogenesis. The soluble factors were collected as conditioned medium (CM) 48 h after culturing stem cells from human exfoliated deciduous teeth (SHED) in serum-free DMEM. SHED-CM significantly increased cell viability of human umbilical vein endothelial cells (HUVECs) in MTT assays and accelerated HUVECs migration in wound healing and Boyden chamber assays. In a Matrigel plug assay of mice, the migrated number of primary endothelial cells was markedly increased in the plug containing SHED-CM or SHED suspension. SHED-CM induced complex tubular structures of HUVECs in a tube formation assay. Furthermore, SHED-CM significantly increased neovascularization from the primary rat aorta, indicating that SHED-CM stimulated primary endothelial cells to promote comprehensive angiogenesis processes. The angiogenic effects of SHED-CM were the same or greater than the effective concentration of VEGF. In conclusion, SHED-CM directly stimulates vascular endothelial cells to promote angiogenesis and is promising for future clinical application.

Eva Mangelsen, Michael Rothe, Angela Schulz et al.

Glomerular hyperfiltration is an important mechanism in the development of albuminuria. During hyperfiltration, podocytes are exposed to increased fluid flow shear stress (FFSS) in Bowman’s space. Elevated Prostaglandin E2 (PGE₂) synthesis and upregulated cyclooxygenase 2 (Cox2) are associated with podocyte injury by FFSS. We aimed to elucidate a PGE₂ autocrine/paracrine pathway in human podocytes (hPC). We developed a modified liquid chromatography tandem mass spectrometry (LC/ESI-MS/MS) protocol to quantify cellular PGE₂, 15-keto-PGE₂, and 13,14-dihydro-15-keto-PGE₂ levels. hPC were treated with PGE₂ with or without separate or combined blockade of prostaglandin E receptors (EP), EP2, and EP4. Furthermore, the effect of FFSS on <i>COX2</i>, <i>PTGER2</i>, and <i>PTGER4</i> expression in hPC was quantified. In hPC, stimulation with PGE₂ led to an EP2- and EP4-dependent increase in cyclic adenosine monophosphate (cAMP) and <i>COX2</i>, and induced cellular PGE₂. <i>PTGER4</i> was downregulated after PGE₂ stimulation in hPC. In the corresponding LC/ESI-MS/MS in vivo analysis at the tissue level, increased PGE₂ and 15-keto-PGE₂ levels were observed in isolated glomeruli obtained from a well-established rat model with glomerular hyperfiltration, the Munich Wistar Frömter rat. <i>COX2</i> and <i>PTGER2</i> were upregulated by FFSS. Our data thus support an autocrine/paracrine COX2/PGE₂ pathway in hPC linked to concerted EP2 and EP4 signaling.

Melissa Dawn Meachem, Julia Bettina Montgomery

Eugenio Leonardo, Ricardo H. Bardales