Bupivacaine in combination with sildenafil (Viagra) and vitamin D3 have anti-inflammatory effects in osteoarthritic chondrocytes

Elisabeth Hansson,Eva Skiöldebrand

doi:10.1016/j.crphar.2021.100066

Abstract

AimsTo treat osteoarthritic chondrocytes and thereby reduce the inflammation with a drug combination that primarily affects 5-HT- and ATP-evoked Ca2+ signaling. In osteoarthritic chondrocytes, Ca2+ signaling is elevated, resulting in increased production of ATP and inflammatory mediators. The expression of TLR4 and Na+/K+-ATPase was used to evaluate the inflammatory status of the cells. Main methodsEquine chondrocytes were collected from joints with mild structural osteoarthritic changes and cultured in monolayers. The cells were treated with a combination of bupivacaine (1 pM) and sildenafil (1 μM) in combination with vitamin D3 (100 nM). A high-throughput screening system, the Flexstation 3 microplate reader, was used to measure intra- and extracellular Ca2+ signaling after exposure to 5-HT, glutamate, or ATP. Expression of inflammatory receptors was assessed by Western blotting. Key findingsDrug treatment substantially reduced 5-HT- and ATP-evoked intracellular Ca2+ release and TLR4 expression compared to those in untreated chondrocytes. The combination of sildenafil, vitamin D3 together with metformin, as the ability to take up glucose is limited, increased Na+/K+-ATPase expression. SignificanceThe combination of these three therapeutic substances at concentrations much lower than usually used, reduced expression of the inflammatory receptor TLR4 and increased the cell membrane enzyme Na+/K+-ATPase, which regulates cell volume and reduces increased intracellular Ca2+ concentrations. These remarkable results indicate that this drug combination has disease-modifying osteoarthritis drug (DMOAD) properties and may be a new clinical therapy for osteoarthritis (OA).

Highlights

Osteoarthritis (OA) is a chronic progressive disease that leads to severe joint pain and the loss of joint mobility in horses and humans (Goldring and Goldring, 2007; van Weeren and de Grauw, 2010)
We previously showed that healthy chondrocytes show an extremely low level compared to OA chondrocytes (Skio€ldebrand et al, 2018, 2019)
Osteoarthritic chondrocytes that had been incubated with LPS for 24 h to further induce an inflammatory response were not further affected by LPS according to their Ca2þ signaling

Summary

Introduction

Osteoarthritis (OA) is a chronic progressive disease that leads to severe joint pain and the loss of joint mobility in horses and humans (Goldring and Goldring, 2007; van Weeren and de Grauw, 2010). No disease-modifying OA drugs (DMOADs) are available (MacKay et al, 2018). Toll-like receptors (TLRs), TLR4, have been identified as potential drug targets for the treatment of inflammatory diseases such as OA (ul Ain et al, 2020). We previously showed that chondrocytes increase their intracellular Ca2þ release, which is associated with increased expression of TLR4 and inflammatory mediators (Skio€ldebrand et al, 2018). The balance between cell-to-cell Ca2þ signaling and extracellular Ca2þ signaling due to increased ATP production and release is changed (Skio€ldebrand et al, 2019; Cotrina et al, 2000; Beamer et al, 2017)

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