scholarly journals Phosphoproteomic analysis of chondrocytes after short-term exposure to inorganic polyphosphate

2021 ◽  
Author(s):  
Uros Kuzmanov ◽  
Rahul Gawri ◽  
Alena Zelinka ◽  
Keith A Russel ◽  
Shin-Haw Lee ◽  
...  
Keyword(s):  
Large Scale ◽  
Articular Chondrocytes ◽  
Treatment Options ◽  
Biological Molecules ◽  
Inorganic Polyphosphate ◽  
Naturally Occurring ◽  
Molecular Events ◽  
Short Term Exposure ◽  
Matrix Production

Osteoarthritis is a debilitating disease of the joint that affects over 230 million people worldwide. Currently there are no treatments that slow the progression of this disease. For these reasons, new biological treatment options are currently being explored. Inorganic polyphosphates are naturally occurring biological molecules that have an anabolic effect on chondrocytes grown in vitro in the presence of Ca2+. We hypothesized that when examining significant changes in protein phosphorylation, key candidates would emerge that could help to elucidate the anabolic effects of polyphosphate on chondrocytes. Therefore, we conducted a large-scale quantitative proteomic and phosphoproteomic study of bovine primary articular chondrocytes after 30-minute treatment with inorganic polyphosphate and Ca2+. Mass spectrometry identified more than 6000 phosphorylation sites on ~1600 chondrocyte phosphoproteins while proteomic analysis detected approximately 4100 proteins. Analysis of the data revealed a swift and dynamic response to polyphosphate after 30 minutes. What emerged from the list of proteins most affected by the treatment were proteins with key roles in chondrogenesis including TNC, IGFBP-5, and CTGF, indicating that polyphosphate plays an important role in chondrocyte metabolism. This phosphoproteome serves as a meaningful resource to help elucidate the molecular events that contribute to extracellular matrix production in cartilage.

scholarly journals Viromers as carriers for mRNA-mediated expression of therapeutic molecules under inflammatory conditions

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Edith Jansig ◽  
Stefanie Geissler ◽  
Vera Rieckmann ◽  
Anja Kuenemund ◽  
Benjamin Hietel ◽  
...  
Keyword(s):  
Treatment Options ◽  
Biological Molecules ◽  
Luciferase Reporter ◽  
Active Components ◽  
Reporter Expression ◽  
Inflammatory Conditions ◽  
Therapeutic Molecules ◽  
In Vitro Protein Expression

Abstract Therapeutic mRNA delivery has been described for several treatment options, such as vaccination and cancer immunotherapy. However, mRNA delivery has to be accompanied by the development and testing of suitable carrier materials due to the instability of mRNAs in human body fluids. In the present study, we investigated the ability of recently developed Viromers to deliver mRNAs in a classical inflammatory setting. We tested mRNAs coding for active components of preclinical (7ND) and approved (sTNF-RII) biologics, in vitro and in vivo. 7ND is an established blocker of the CCR2 axis, whereas sTNF-RII is the active component of the approved drug Etanercept. Viromer/mRNA complexes were transfected into murine macrophages in vitro. Protein expression was analysed using Luciferase reporter expression and mainly identified in spleen, blood and bone marrow in vivo. 7ND-mRNA delivery led to efficient blockage of monocytes infiltration in thioglycolate-induced peritonitis in mice, underlining the ability of Viromers to deliver a therapeutic mRNA cargo without overt toxicity. Therefore, we propose Viromer-based mRNA delivery as a suitable option for the treatment of inflammatory disorders beyond infusion of biological molecules.

In vitro effects of meloxicam on metabolism in articular chondrocytes from dogs with naturally occurring osteoarthritis

2013 ◽  
Vol 74 (9) ◽  
pp. 1198-1205 ◽  
Author(s):  
Steven C. Budsberg ◽  
Aaron M. Stoker ◽  
Spencer A. Johnston ◽  
William Liska ◽  
Lisa R. Reno ◽  
...  
Keyword(s):  
Articular Chondrocytes ◽  
Naturally Occurring ◽  
In Vitro Effects

scholarly journals ST3GAL1 is a target of the SOX2-GLI1 transcriptional complex and promotes melanoma metastasis through AXL

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Silvia Pietrobono ◽  
Giulia Anichini ◽  
Cesare Sala ◽  
Fabrizio Manetti ◽  
Luciana L. Almada ◽  
...  
Keyword(s):  
Therapeutic Potential ◽  
Treatment Options ◽  
Blood Stream ◽  
In Vivo Studies ◽  
Melanoma Metastasis ◽  
Melanoma Invasion ◽  
Molecular Events ◽  
Transcriptional Complex

AbstractUnderstanding the molecular events controlling melanoma progression is of paramount importance for the development of alternative treatment options for this devastating disease. Here we report a mechanism regulated by the oncogenic SOX2-GLI1 transcriptional complex driving melanoma invasion through the induction of the sialyltransferase ST3GAL1. Using in vitro and in vivo studies, we demonstrate that ST3GAL1 drives melanoma metastasis. Silencing of this enzyme suppresses melanoma invasion and significantly reduces the ability of aggressive melanoma cells to enter the blood stream, colonize distal organs, seed and survive in the metastatic environment. Analysis of glycosylated proteins reveals that the receptor tyrosine kinase AXL is a major effector of ST3GAL1 pro-invasive function. ST3GAL1 induces AXL dimerization and activation that, in turn, promotes melanoma invasion. Our data support a key role of the ST3GAL1-AXL axis as driver of melanoma metastasis, and highlight the therapeutic potential of targeting this axis to treat metastatic melanoma.

scholarly journals Single Agent Polysaccharopeptide Delays Metastases and Improves Survival in Naturally Occurring Hemangiosarcoma

2012 ◽  
Vol 2012 ◽  
pp. 1-8 ◽  
Author(s):  
Dorothy Cimino Brown ◽  
Jennifer Reetz
Keyword(s):  
Cancer Patients ◽  
Large Scale ◽  
Single Agent ◽  
Treatment Strategies ◽  
World Health ◽  
High Dose ◽  
Survival Times ◽  
Double Blind ◽  
Naturally Occurring

The 2008 World Health Organization World Cancer Report describes global cancer incidence soaring with many patients living in countries that lack resources for cancer control. Alternative treatment strategies that can reduce the global disease burden at manageable costs must be developed. Polysaccharopeptide (PSP) is the bioactive agent from the mushroomCoriolus versicolor. Studies indicate PSP has in vitro antitumor activities and inhibits the growth of induced tumors in animal models. Clear evidence of clinically relevant benefits of PSP in cancer patients, however, is lacking. The investment of resources required to complete large-scale, randomized controlled trials of PSP in cancer patients is more easily justified if antitumor and survival benefits are documented in a complex animal model of a naturally occurring cancer that parallels human disease. Because of its high metastatic rate and vascular origin, canine hemangiosarcoma is used for investigations in antimetastatic and antiangiogenic therapies. In this double-blind randomized multidose pilot study, high-dose PSP significantly delayed the progression of metastases and afforded the longest survival times reported in canine hemangiosarcoma. These data suggest that, for those cancer patients for whom advanced treatments are not accessible, PSP as a single agent might offer significant improvements in morbidity and mortality.

Proper expression of helix–loop–helix protein Id2 is important to chondrogenic differentiation of ATDC5 cells

2009 ◽  
Vol 419 (3) ◽  
pp. 635-643 ◽  
Author(s):  
Liu Yang ◽  
Xiaoyun Ma ◽  
Anne Lyone ◽  
Junhui Zou ◽  
Michael L. Blackburn ◽  
...  
Keyword(s):  
Chondrogenic Differentiation ◽  
Large Scale ◽  
Articular Chondrocytes ◽  
Type Ii Collagen ◽  
Inverse Pcr ◽  
Pcr Analysis ◽  
Insertion Mutagenesis ◽  
Chondroprogenitor Cells

The process of chondrogenesis can be mimicked in vitro by insulin treatment of mouse ATDC5 chondroprogenitor cells. To identify novel factors that are involved in the control of chondrogenesis, we carried out a large-scale screening through retroviral insertion mutagenesis and isolated a fast-growing ATDC5 clone incapable of chondrogenic differentiation. Inverse-PCR analysis of this clone revealed that the retroviral DNA was inserted into the promoter region of mouse Id2 (inhibitor of DNA-binding protein 2) gene. This retroviral insertion increased Id2 protein levels to twice those found in normal ATDC5 cells. To investigate whether an elevated level of Id2 protein was responsible for inhibition of chondrogenic differentiation, ATDC5 cells were infected with a retrovirus to stably express Id2. ATDC5 cells expressing ectopic Id2 exhibited signs of de-differentiation, such as rapid growth, and insulin failed to induce expression of Sox9 (Sry-type high-mobility-group box 9) or matrix genes such as type II collagen (COL2) in these cells. When endogenous Id2 was knocked down by siRNA (small interfering RNA) in ATDC5 cells, expression of Sox9 and COL2 was increased and chondrogenic differentiation was accelerated. To examine how Id2 is expressed in chondrocytes in vivo, we carried out immunostaining of E16.5 mouse embryos and found that Id2 is expressed in articular chondrocytes and proliferating chondrocytes, but barely detectable in hypertrophic chondrocytes. Our results suggest that proper expression of Id2 is important to achieving a fine balance between growth and differentiation during chondrogenesis.

scholarly journals Mildred Rebstock: Profile of the Medicinal Chemist Who Synthesized Chloramphenicol

2019 ◽  
Vol 63 (6) ◽  
Author(s):  
David M. Aronoff
Keyword(s):  
United States ◽  
Large Scale ◽  
The United States ◽  
70Th Anniversary ◽  
Scale Production ◽  
Naturally Occurring ◽  
Large Scale Production

ABSTRACTThis year marks the 70th anniversary since Parke-Davis and Company announced the synthesis of chloramphenicol, the first naturally occurring antibiotic to be chemically generatedin vitrofor large-scale production. The effort was led by the chemist Mildred Rebstock, Ph.D., (1919 to 2011), who would turn 100 years old this year. Her accomplishment, at a time when very few chemists in the United States were women, was celebrated internationally. This commentary reviews her important contribution.

scholarly journals Physioxia Has a Beneficial Effect on Cartilage Matrix Production in Interleukin-1 Beta-Inhibited Mesenchymal Stem Cell Chondrogenesis

Cells ◽  
2019 ◽  
Vol 8 (8) ◽  
pp. 936 ◽  
Author(s):  
Girish Pattappa ◽  
Ruth Schewior ◽  
Isabelle Hofmeister ◽  
Jennifer Seja ◽  
Johannes Zellner ◽  
...  
Keyword(s):  
Articular Cartilage ◽  
Beneficial Effect ◽  
Articular Chondrocytes ◽  
Interleukin 1 ◽  
Cartilage Matrix ◽  
Interleukin 1Β ◽  
Low Oxygen ◽  
Matrix Gene ◽  
Matrix Production

Osteoarthritis (OA) is a degenerative condition that involves the production of inflammatory cytokines (e.g., interleukin-1β (IL-1β), tumour necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6)) that stimulate degradative enzymes, matrix metalloproteinases (MMPs) and aggrecanases (ADAMTS) resulting in articular cartilage breakdown. The presence of interleukin-1β (IL-1β) is one reason for poor clinical outcomes in current cell-based tissue engineering strategies for treating focal early osteoarthritic defects. Mesenchymal stem cells (MSCs) are a potential cell source for articular cartilage regeneration, although IL-1β has been shown to inhibit in vitro chondrogenesis. In vivo, articular chondrocytes reside under a low oxygen environment between 2–5% oxygen (physioxia) and have been shown to enhance in vitro MSC chondrogenic matrix content with reduced hypertrophic marker expression under these conditions. The present investigation sought to understand the effect of physioxia on IL-1β inhibited MSC chondrogenesis. MSCs expanded under physioxic (2% oxygen) and hyperoxic (20%) conditions, then chondrogenically differentiated as pellets in the presence of TGF-β1 and either 0.1 or 0.5 ng/mL IL-1β. Results showed that there were donor variations in response to physioxic culture based on intrinsic GAG content under hyperoxia. In physioxia responsive donors, MSC chondrogenesis significantly increased GAG and collagen II content, whilst hypertrophic markers were reduced compared with hyperoxia. In the presence of IL-1β, these donors showed a significant increase in cartilage matrix gene expression and GAG content relative to hyperoxic conditions. In contrast, a set of MSC donors were unresponsive to physioxia and showed no significant increase in matrix production independent of IL-1β presence. Thus, physioxia has a beneficial effect on MSC cartilage matrix production in responsive donors with or without IL-1β application. The mechanisms controlling the MSC chondrogenic response in both physioxia responsive and unresponsive donors are to be elucidated in future investigations.

scholarly journals Production and Purification of Artificial Circular RNA Sponges for Application in Molecular Biology and Medicine

2020 ◽  
Vol 3 (2) ◽  
pp. 42 ◽  
Author(s):  
Janina Breuer ◽  
Oliver Rossbach
Keyword(s):  
Molecular Biology ◽  
Large Scale ◽  
Circular Rna ◽  
In Vitro Transcription ◽  
Circular Rnas ◽  
Small Scale ◽  
Limiting Factor ◽  
Scale Production ◽  
Naturally Occurring

Characterized by their covalently closed structure and thus an elevated stability compared to linear RNA molecules, circular RNAs (circRNAs) form a novel class of mainly non-coding RNAs. Although the biological functions of naturally occurring circRNAs are largely unknown, they were reported to act as molecular sponges, sequestering microRNAs (miRNAs), resulting in a de-repression of target mRNAs. Taking these characteristics of naturally occurring circRNAs into account, artificial circRNAs could be a potential tool in molecular biology and medicine. Using the Hepatitis C virus (HCV) as a model system, this application of artificial circular RNAs was demonstrated. The virus requires cellular miRNA miR-122 for its life cycle, and circRNAs specifically engineered to efficiently sequester this miRNA impacted viral propagation. Since in this context the production of engineered circRNA remains the limiting factor, we present a method to produce and efficiently purify artificial circRNA sponges (ciRS) in vitro. In this protocol we provide insights into a small-scale and large-scale production technique of artificial circular RNA sponges relying on in vitro transcription and RNA ligation.

scholarly journals The NF-κB transcriptional footprint is essential for SARS-CoV-2 replication

2021 ◽  
Author(s):  
Benjamin E. Nilsson-Payant ◽  
Skyler Uhl ◽  
Adrien Grimont ◽  
Ashley S. Doane ◽  
Phillip Cohen ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Large Scale ◽  
Cytokine Production ◽  
Antiviral Treatment ◽  
Treatment Options ◽  
Type I ◽  
Mortality And Morbidity ◽  
The World ◽  
Infected Cells

SARS-CoV-2, the etiological agent of COVID-19, is characterized by a delay in Type I interferon (IFN-I)-mediated antiviral defenses alongside robust cytokine production. Here we investigate the underlying molecular basis for this imbalance and implicate virus-mediated activation of NF-κB in the absence of other canonical IFN-I-related transcription factors. Epigenetic and single cell transcriptomic analyses show a selective NF-κB signature that was most prominent in infected cells. Disruption of NF-κB signaling through the silencing of the NF-κB transcription factors p65 or p50 resulted in loss of virus replication that was rescued upon reconstitution. These findings could be further corroborated with the use of NF-κB inhibitors, which reduced SARS-CoV-2 replication in vitro . These data suggest that the robust cytokine production in response to SARS-CoV-2, despite a diminished IFN-I response, is the product of a dependency on NF-κB for viral replication. IMPORTANCE The COVID-19 pandemic has caused significant mortality and morbidity around the world. Although effective vaccines have been developed, large parts of the world remain unvaccinated while new SARS-CoV-2 strains keep emerging. Furthermore, despite extensive efforts and large-scale drug screenings, no fully effective antiviral treatment options have been discovered yet. Therefore, it is of the utmost importance to gain a better understanding of essential factors driving SARS-CoV-2 replication in order to be able to develop novel approaches to target SARS-CoV-2 biology.

scholarly journals Encouraging Early Outcomes of Treatment With Arsenic Trioxide Combined With Chemotherapy for Alveolar Rhabdomyosarcoma in Children: 4 Case Reports

2021 ◽  
Vol 11 ◽  
Author(s):  
Xiaomin Peng ◽  
Xilin Xiong ◽  
Yang Li ◽  
Chuchu Feng ◽  
Hongyan Liu ◽  
...  
Keyword(s):  
Arsenic Trioxide ◽  
Large Scale ◽  
Case Reports ◽  
Treatment Options ◽  
Late Toxicity ◽  
Alveolar Rhabdomyosarcoma ◽  
Combined Chemotherapy ◽  
Chemotherapeutic Regimen ◽  
Comprehensive Management

BackgroundAlveolar rhabdomyosarcoma (ARMS) is a subtype of rhabdomyosarcoma characterized by its aggressive behavior and poor prognosis, highlighting the need for novel treatment options. Arsenic trioxide (ATO) has been shown to specifically inhibit tumor growth and the metastasis of ARMS in vitro by acting on the hedgehog pathway. Here we report on a pilot clinical study to evaluate the activity of an ATO-combined chemotherapy approach for the treatment of ARMS patients.MethodsWe designed a therapeutic schedule of an ATO-combined chemotherapy, incorporating comprehensive management according to the Intergroup Rhabdomyosarcoma Study Group protocol. ATO was administered at 0.16 mg/kg per day over 8 h via an IV for 10 days combined with a chemotherapeutic regimen of vincristine, actinomycin, and cyclophosphamide (VAC regimen) on the third day, which was repeated every 21 days. A total of eight cycles of ATO-combined chemotherapy were applied throughout the entire scheme.ResultsA total of three refractory/recurrent and one untreated ARMS patient, three male and one female, with a median age of 5.8 years (range, 5.1 to 12.5 years), were enrolled in the present study. All patients were sensitive to combined chemotherapy with ATO and achieved partial or complete remission during treatment. Except for reversible gastrointestinal reaction and myelosuppression, no other adverse events were observed during the process of treatment.ConclusionsThe combined chemotherapy of ATO and the VAC regimen exhibited beneficial activities against ARMS in pediatrics and was well tolerated, but prospective large-scale clinical trials are warranted to determine the long-term efficacy, optimal courses, and late toxicity in this population.

Sign in / Sign up

Export Citation Format

Share Document