Development of Glycerol-Rose Bengal-Polidocanol (GRP) foam for enhanced sclerosis of a cyst for cystic diseases

Polycystic kidney disease (PKD) is a common genetic disorder that results in a proliferating and enlarging cyst and ultimately leads to loss of kidney function. Because an enlarged cyst is a primary factor for limited kidney function, the large cyst is surgically removed by laparoscopic deroofing or sclerosant. This a relatively nascent treatment method entails complications and sometimes fail due to the cyst fluid refilling and infection. This study proposes using a more stable and effective polidocanol foam with glycerol and Rose Bengal (GRP form) to prevent cyst regeneration and irritation, which is caused by the required body movement during the treatment. Specifically, the foam retention time and viscosity were increased by adding glycerol up to 10% (w/v). The GRP form inhibited cellular proliferation and disrupted cellular junctions, e-cadherin, and cyst formation, demonstrated by the LDH, Live and Dead, and re-plating culture assays. The GRP foam was shown to be a safe and effective treatment as a commercial grade polidocanol foam form by an in vivo study in which subcutaneously injected mice injected with commercial 3% polidocanol, and the GRP foam showed no difference in inflammation. Thus, this study provides an advanced polidocanol form by adding glycerol and Rose-Bengal to help existing sclerotherapy.

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In vitro 3D phenotypic drug screen identifies celastrol as an effective in vivo inhibitor of polycystic kidney disease

Journal of Molecular Cell Biology ◽

10.1093/jmcb/mjz029 ◽

2019 ◽

Vol 12 (8) ◽

pp. 644-653 ◽

Cited By ~ 2

Author(s):

Tijmen H Booij ◽

Wouter N Leonhard ◽

Hester Bange ◽

Kuan Yan ◽

Michiel Fokkelman ◽

...

Keyword(s):

Kidney Disease ◽

Polycystic Kidney Disease ◽

Genetic Disorder ◽

Screening Method ◽

Renal Cysts ◽

Cyst Formation ◽

Polycystic Kidney ◽

Cyst Growth

Abstract Polycystic kidney disease (PKD) is a prevalent genetic disorder, characterized by the formation of kidney cysts that progressively lead to kidney failure. The currently available drug tolvaptan is not well tolerated by all patients and there remains a strong need for alternative treatments. The signaling rewiring in PKD that drives cyst formation is highly complex and not fully understood. As a consequence, the effects of drugs are sometimes difficult to predict. We previously established a high throughput microscopy phenotypic screening method for quantitative assessment of renal cyst growth. Here, we applied this 3D cyst growth phenotypic assay and screened 2320 small drug-like molecules, including approved drugs. We identified 81 active molecules that inhibit cyst growth. Multi-parametric phenotypic profiling of the effects on 3D cultured cysts discriminated molecules that showed preferred pharmacological effects above genuine toxicological properties. Celastrol, a triterpenoid from Tripterygium Wilfordii, was identified as a potent inhibitor of cyst growth in vitro. In an in vivo iKspCre-Pkd1lox,lox mouse model for PKD, celastrol inhibited the growth of renal cysts and maintained kidney function.

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In Vivo Assessment of Metabolic Abnormality in Alport Syndrome Using Hyperpolarized [1-13C] Pyruvate MR Spectroscopic Imaging

Metabolites ◽

10.3390/metabo11040222 ◽

2021 ◽

Vol 11 (4) ◽

pp. 222

Author(s):

Nguyen-Trong Nguyen ◽

Eun-Hui Bae ◽

Luu-Ngoc Do ◽

Tien-Anh Nguyen ◽

Ilwoo Park ◽

...

Keyword(s):

Disease Progression ◽

Alport Syndrome ◽

Genetic Disorder ◽

Lactate Production ◽

Clinical Importance ◽

Metabolic Imaging ◽

Renal Metabolism ◽

Hyperpolarized 13C ◽

And Control

Alport Syndrome (AS) is a genetic disorder characterized by impaired kidney function. The development of a noninvasive tool for early diagnosis and monitoring of renal function during disease progression is of clinical importance. Hyperpolarized 13C MRI is an emerging technique that enables non-invasive, real-time measurement of in vivo metabolism. This study aimed to investigate the feasibility of using this technique for assessing changes in renal metabolism in the mouse model of AS. Mice with AS demonstrated a significant reduction in the level of lactate from 4- to 7-week-old, while the levels of lactate were unchanged in the control mice over time. This reduction in lactate production in the AS group accompanied a significant increase of PEPCK expression levels, indicating that the disease progression in AS triggered the gluconeogenic pathway and might have resulted in a decreased lactate pool size and a subsequent reduction in pyruvate-to-lactate conversion. Additional metabolic imaging parameters, including the level of lactate and pyruvate, were found to be different between the AS and control groups. These preliminary results suggest that hyperpolarized 13C MRI might provide a potential noninvasive tool for the characterization of disease progression in AS.

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Study Design and Baseline Characteristics of the CARDINAL Trial: A Phase 3 Study of Bardoxolone Methyl in Patients with Alport Syndrome

American Journal of Nephrology ◽

10.1159/000513777 ◽

2021 ◽

pp. 1-10

Author(s):

Glenn M. Chertow ◽

Gerald B. Appel ◽

Sharon Andreoli ◽

Sripal Bangalore ◽

Geoffrey A. Block ◽

...

Keyword(s):

Kidney Function ◽

Alport Syndrome ◽

Genetic Disorder ◽

Geometric Mean ◽

Genetic Diagnosis ◽

Significant Loss ◽

The European Union ◽

Phase 3 ◽

Double Blind ◽

Histologic Diagnosis

Introduction: Alport syndrome is a rare genetic disorder that affects as many as 60,000 persons in the USA and a total of 103,000 persons (<5 per 10,000) in the European Union [1, 2]. It is the second most common inherited cause of kidney failure and is characterized by progressive loss of kidney function that often leads to end-stage kidney disease. Currently, there are no approved disease-specific agents for therapeutic use. We designed a phase 3 study (CARDINAL; NCT03019185) to evaluate the safety, tolerability, and efficacy of bardoxolone methyl in patients with Alport syndrome. Methods: The CARDINAL phase 3 study is an international, multicenter, double-blind, placebo-controlled, randomized registrational trial. Eligible patients were of ages 12–70 years with confirmed genetic or histologic diagnosis of Alport syndrome, eGFR 30–90 mL/min/1.73 m2, and urinary albumin to creatinine ratio (UACR) ≤3,500 mg/g. Patients with B-type natriuretic peptide values >200 pg/mL at baseline or with significant cardiovascular histories were excluded. Patients were randomized 1:1 to bardoxolone methyl or placebo, with stratification by baseline UACR. Results: A total of 371 patients were screened, and 157 patients were randomly assigned to receive bardoxolone methyl (n = 77) or placebo (n = 80). The average age at screening was 39.2 years, and 23 (15%) were <18 years of age. Of the randomized population, 146 (93%) had confirmed genetic diagnosis of Alport syndrome, and 62% of patients had X-linked mode of inheritance. Mean baseline eGFR was 62.7 mL/min/1.73 m2, and the geometric mean UACR was 141.0 mg/g. The average annual rate of eGFR decline prior to enrollment in the study was −4.9 mL/min/1.73 m2 despite 78% of the patient population receiving ACE inhibitor (ACEi) or ARB therapy. Discussion/Conclusion: CARDINAL is one of the largest interventional, randomized controlled trials in Alport syndrome conducted to date. Despite the use of ACEi or ARB, patients were experiencing significant loss of kidney function prior to study entry.

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RAE-1 ligands for the NKG2D receptor are regulated by E2F transcription factors, which control cell cycle entry

Journal of Experimental Medicine ◽

10.1084/jem.20120565 ◽

2012 ◽

Vol 209 (13) ◽

pp. 2409-2422 ◽

Cited By ~ 77

Author(s):

Heiyoun Jung ◽

Benjamin Hsiung ◽

Kathleen Pestal ◽

Emily Procyk ◽

David H. Raulet

Keyword(s):

Cell Cycle ◽

Transcription Factors ◽

Stress Responses ◽

Transcriptional Activation ◽

Posttranscriptional Regulation ◽

Cellular Proliferation ◽

Control Cell ◽

T Cell Subsets ◽

Cell Cycle Entry

The NKG2D stimulatory receptor expressed by natural killer cells and T cell subsets recognizes cell surface ligands that are induced on transformed and infected cells and facilitate immune rejection of tumor cells. We demonstrate that expression of retinoic acid early inducible gene 1 (RAE-1) family NKG2D ligands in cancer cell lines and proliferating normal cells is coupled directly to cell cycle regulation. Raet1 genes are directly transcriptionally activated by E2F family transcription factors, which play a central role in regulating cell cycle entry. Induction of RAE-1 occurred in primary cell cultures, embryonic brain cells in vivo, and cells in healing skin wounds and, accordingly, wound healing was delayed in mice lacking NKG2D. Transcriptional activation by E2Fs is likely coordinated with posttranscriptional regulation by other stress responses. These findings suggest that cellular proliferation, as occurs in cancer cells but also other pathological conditions, is a key signal tied to immune reactions mediated by NKG2D-bearing lymphocytes.

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UV-induced Hyperphosphorylation of Replication Protein A Depends on DNA Replication and Expression of ATM Protein

Molecular Biology of the Cell ◽

10.1091/mbc.12.5.1199 ◽

2001 ◽

Vol 12 (5) ◽

pp. 1199-1213 ◽

Cited By ~ 58

Author(s):

Gregory G. Oakley ◽

Lisa I. Loberg ◽

Jiaqin Yao ◽

Mary A. Risinger ◽

Remy L. Yunker ◽

...

Keyword(s):

Dna Replication ◽

Uv Radiation ◽

Excision Repair ◽

Genetic Disorder ◽

Protein A ◽

Dna Recombination ◽

Delayed Response ◽

Replication Intermediates

Exposure to DNA-damaging agents triggers signal transduction pathways that are thought to play a role in maintenance of genomic stability. A key protein in the cellular processes of nucleotide excision repair, DNA recombination, and DNA double-strand break repair is the single-stranded DNA binding protein, RPA. We showed previously that the p34 subunit of RPA becomes hyperphosphorylated as a delayed response (4–8 h) to UV radiation (10–30 J/m2). Here we show that UV-induced RPA-p34 hyperphosphorylation depends on expression of ATM, the product of the gene mutated in the human genetic disorder ataxia telangiectasia (A-T). UV-induced RPA-p34 hyperphosphorylation was not observed in A-T cells, but this response was restored by ATM expression. Furthermore, purified ATM kinase phosphorylates the p34 subunit of RPA complex in vitro at many of the same sites that are phosphorylated in vivo after UV radiation. Induction of this DNA damage response was also dependent on DNA replication; inhibition of DNA replication by aphidicolin prevented induction of RPA-p34 hyperphosphorylation by UV radiation. We postulate that this pathway is triggered by the accumulation of aberrant DNA replication intermediates, resulting from DNA replication fork blockage by UV photoproducts. Further, we suggest that RPA-p34 is hyperphosphorylated as a participant in the recombinational postreplication repair of these replication products. Successful resolution of these replication intermediates reduces the accumulation of chromosomal aberrations that would otherwise occur as a consequence of UV radiation.

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Antagonism of endogenous putative P2Y receptors reduces the growth of MDCK-derived cysts cultured in vitro

AJP Renal Physiology ◽

10.1152/ajprenal.00103.2006 ◽

2007 ◽

Vol 292 (1) ◽

pp. F15-F25 ◽

Cited By ~ 25

Author(s):

Clare M. Turner ◽

Brian F. King ◽

Kaila S. Srai ◽

Robert J. Unwin

Keyword(s):

Therapeutic Potential ◽

Cyst Formation ◽

P2y Receptors ◽

P2x Receptor ◽

P2y Receptor ◽

Cyst Size ◽

Reactive Blue 2 ◽

Cyst Growth

P2Y receptors couple to G proteins and either mobilize intracellular Ca2+ or alter cAMP levels to modulate the activity of Ca2+- and cAMP-sensitive ion channels. We hypothesize that increased ion transport into the lumen of MDCK cysts can osmotically drive fluid movement and increase cyst size. Furthermore, activation of the adenylate cyclase/cAMP pathway may trigger cell proliferation via an extracellular signal-related kinase cascade. To test this hypothesis, several P2Y receptor inhibitors were used on the MDCK in vitro model of renal cyst formation. The nonspecific P2 receptor inhibitors reactive blue 2 and suramin reduced cyst growth significantly, as did PPADS and, to a lesser extent, the P2Y1-specific antagonist MRS2179. Cyst growth was reduced by ∼50% when ATP was removed from the culture medium with apyrase, although stable analogs of ATP failed to increase cyst size. The nonselective P2X receptor inhibitor Coomassie brilliant blue G was ineffective at reducing cyst growth, suggesting no involvement of P2X receptors. Finally, the presence of selective inhibitors of ERK activation (either PD98059 or U0126) greatly reduced cyst growth, whereas in untreated cysts ERK activity was observed to increase with time. We conclude that stimulation of endogenous P2Y receptors by extracellular ATP increases growth of MDCK cysts via cAMP-dependent activation of the ERK pathway. P2Y receptor antagonists may have therapeutic potential in reducing cyst size and slowing disease progression; although further studies in vitro and in vivo are needed to investigate the specificity and role of these P2Y receptors in renal cystic diseases.

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The c-myc proto-oncogene regulates cardiac development in transgenic mice.

Molecular and Cellular Biology ◽

10.1128/mcb.10.7.3709 ◽

1990 ◽

Vol 10 (7) ◽

pp. 3709-3716 ◽

Cited By ~ 128

Author(s):

T Jackson ◽

M F Allard ◽

C M Sreenan ◽

L K Doss ◽

S P Bishop ◽

...

Keyword(s):

Transgenic Mice ◽

Mrna Expression ◽

Cardiac Myocyte ◽

Cellular Proliferation ◽

Cardiac Development ◽

Constitutive Expression ◽

Transgenic Animals ◽

Hypertrophic Growth ◽

Myocyte Proliferation

During the maturation of the cardiac myocyte, a transition occurs from hyperplastic to hypertrophic growth. The factors that control this transition in the developing heart are unknown. Proto-oncogenes such as c-myc have been implicated in the regulation of cellular proliferation and differentiation, and in the heart the switch from myocyte proliferation to terminal differentiation is synchronous with a decrease in c-myc mRNA abundance. To determine whether c-myc can influence myocyte proliferation or differentiation, we examined the in vivo effect of increasing c-myc expression during embryogenesis and of preventing the decrease in c-myc mRNA expression that normally occurs during cardiac development. The model system used was a strain of transgenic mice exhibiting constitutive expression of c-myc mRNA in cardiac myocytes throughout development. In these transgenic mice, increased c-myc mRNA expression was found to be associated with both atrial and ventricular enlargement. This increase in cardiac mass was secondary to myocyte hyperplasia, with the transgenic hearts containing more than twice as many myocytes as did nontransgenic hearts. The results suggest that in the transgenic animals there is additional hyperplastic growth during fetal development. However, this additional proliferative growth is not reflected in abnormal myocyte maturation, as assessed by the expression of the cardiac and skeletal isoforms of alpha-actin. The results of this study indicate that constitutive expression of c-myc mRNA in the heart during development results in enhanced hyperplastic growth and suggest a regulatory role for this proto-oncogene in cardiac myogenesis.

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Constitutive and Regulated Shedding of Soluble FGF Receptors Releases Biologically Active Inhibitors of FGF-2

International Journal of Molecular Sciences ◽

10.3390/ijms22052712 ◽

2021 ◽

Vol 22 (5) ◽

pp. 2712

Author(s):

Anne Hanneken ◽

Maluz Mercado ◽

Pamela Maher

Keyword(s):

Cell Surface ◽

Ligand Binding ◽

Cellular Proliferation ◽

Biological Activities ◽

Biological Properties ◽

Matrix Metalloproteases ◽

Biologically Active ◽

Ectodomain Shedding ◽

High Affinity

The identification of soluble fibroblast growth factor (FGF) receptors in blood and the extracellular matrix has led to the prediction that these proteins modulate the diverse biological activities of the FGF family of ligands in vivo. A recent structural characterization of the soluble FGF receptors revealed that they are primarily generated by proteolytic cleavage of the FGFR-1 ectodomain. Efforts to examine their biological properties are now focused on understanding the functional consequences of FGFR-1 ectodomain shedding and how the shedding event is regulated. We have purified an FGFR-1 ectodomain that is constitutively cleaved from the full-length FGFR-1(IIIc) receptor and released into conditioned media. This shed receptor binds FGF-2; inhibits FGF-2-induced cellular proliferation; and competes with high affinity, cell surface FGF receptors for ligand binding. FGFR-1 ectodomain shedding downregulates the number of high affinity receptors from the cell surface. The shedding mechanism is regulated by ligand binding and by activators of PKC, and the two signaling pathways appear to be independent of each other. Deletions and substitutions at the proposed cleavage site of FGFR-1 do not prevent ectodomain shedding. Broad spectrum inhibitors of matrix metalloproteases decrease FGFR-1 ectodomain shedding, suggesting that the enzyme responsible for constitutive, ligand-activated, and protein kinase C-activated shedding is a matrix metalloprotease. In summary, shedding of the FGFR-1 ectodomain is a highly regulated event, sharing many features with a common system that governs the release of diverse membrane proteins from the cell surface. Most importantly, the FGFR ectodomains are biologically active after shedding and are capable of functioning as inhibitors of FGF-2.

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GENOTOXIC AND CARCINOGENIC STUDIES OF NORGESTREL IN DROSOPHILA MELANOGASTER

Asian Journal of Pharmaceutical and Clinical Research ◽

10.22159/ajpcr.2018.v11i10.27374 ◽

2018 ◽

Vol 11 (10) ◽

pp. 372

Author(s):

Abou-eisha A ◽

Adel E El-din

Keyword(s):

Somatic Mutation ◽

Cellular Proliferation ◽

Research Work ◽

The Body ◽

The Other ◽

Direct Stimulation ◽

Carcinogenic Activity ◽

Genetic Examination ◽

First Time

Objective: The aim of this study was to investigate, for the first time, the possible in vivo genotoxic and carcinogenic activity associated with exposure to norgestrel (NGT) drug through employing the very recently established and adjusted genotoxic and tumorigenic methods in Drosophila melanogaster.Methods: Two in vivo genotoxic test systems were used; one detects the somatic mutation and recombination effects (somatic mutation and recombination test [SMART] wing-spot test) and the other detects the primary DNA damage (the comet test) in the body cells of D. melanogaster. On the other hand, the warts (wts)-based SMART assay is a vital genetic examination in Drosophila used to identify and characterize cancer potential of compounds.Results: Four experimental doses of NGT were used (ranging from 0.24 μM to 16 μM). NGT was found to be non-genotoxic at all tested concentrations even at the highest dose level 16 μM and failed to increase the frequency of tumors in the somatic cells of D. melanogaster.Conclusion: Our results strengthen the hypothesis that steroidal drugs might act through a non-genotoxic carcinogen mechanism where the carcinogenic properties occur by direct stimulation of cellular proliferation through a steroid receptor-mediated mechanism. In addition, the results obtained in this research work may contribute to highlighting the importance of NGT as a potent neuroprotective antioxidant drug.

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