Prilocaine induces apoptosis in osteoblastic cells

Abstract

Purpose: To determine whether prilocaine, a local anesthetic, induces apoptosis in osteoblastic cells. Methods: After reaching subconfluence, human osteoblastic Saos-2 and HG63 cells and mouse osteoblastic MC3T3-E I cells were exposed for 48 hr to varying concentrations of prilocaine up to I 0 mM and the cytotoxicity of the cells was analyzed by phase-contrast microscopy and WST-I assay. Saos-2 cells treated for 48 hr with 5 mM prilocaine were stained with Hoechst 33342 and nuclear fragmentation was examined under a fluorescence microscope. DNA was extracted from the cells treated with 5 mM prilocaine and DNA ladder formation (a hallmark of apoptosis) was analyzed by agarose gel electrophoresis. Rr.s~t: Prilocaine induced cell death in Saos-2 cells in a dose-and time-dependent manner up to the concentration of 10 mM. Marked nuclear condensation and fragmentation of chromatin were observed in the prilocainetreated cells. DNA ladder formation also was induced by prilocaine treatment, Prilocaine-induced DNA ladder formation was dose-dependent with maximal effect at a concentration of 5 mM and was time-dependent from 12 to 48 hr. DNA ladder formation was also induced by prilocaine treatment in human osteoblastic MG63 cells and mouse osteoblastic MC3T3-EI cells. Cycloheximide prevented prilocaine-induced apoptosis in Saos-2 cells in a dose-dependent fashion up to 20/IM as determined by WST-I assay and DNA ladder formation in agarose gel electrophoresis. ConckLsion: Osteoblastic cells treated with prilocaine exhibit both morphological and biochemical features indicative of apoptosis. The apoptotic mechanisms involve transcriptional regulation of specific proteins or protein synthesis.Objectif: D&errniner si la prilocaYne, un anesth&ique local, a induit rapoptose de cellules ost~oblastiques. M~.thode : Apt& avoir atteint le stade de sous-confluence, des cellules ostEoblastiques humaines Saos-2 et MG63 et ost~oblastiques de souris HC3T3-E I ont &~ expos~es pendant 48 h ~ des concentrations variables de prilocaYne allant jusqu'~ I0 mM et la cytotoxicit~ des cellules a &~ analys~e par microscopie en contraste de phase et par dosage WST-I. Les cellules Saos-2 traitEes pendant 48 h avec 5 mM de prilocaYne ont Et~ color~es avec le Hoechst 33342 et la fragmentation nuclEaire a Et~ examinee sous microscopie en fluorescence. I'ADN a ~t~ extrait et la formation if&helle d'ADN (signe cardinal de l'apoptose) a EtE analys~e par Electrophor&e sur gel d'agarose. ]~.~talm 9 La prilocaine a induit la mort des cellules Saos-2 d'une mani&e d~pendante de la dose et du temps jusqu'~ une concentration de I0 mM. En effet, la prilocaYne a induit dans les cellules trait&s une condensation nucl~aire marquee et la fragmentation de chromatine de m~me que la formation if&helle d'ADN. La formation d'ADN, d~pendante de la dose, a connu son effet maximal avec une concentration de 5 mH et ~tait d~pendante du temps entre 12 et 48 h. La formation d'ADN a Et~ aussi induite par la prilocaYne clans les cellules humaines MG63 et dans les cellules de souris ...

Ectopic expression of clusterin/apolipoprotein J or Bcl-2 decreases the sensitivity of HaCaT cells to toxic effects of ropivacaine

“…However, bupivacaine was shown to have a more potent cytotoxic effect than ropivacaine on the same cell types [6]. Other local anesthetics appear to have similar effects on various cell types [9][10][11][12][13][14]. Prilocaine was found to induce apoptosis in mouse and human osteoblastic cells [11] and lidocaine was shown to inhibit the proliferation of normal human fibroblasts, endothelial cells and keratinocytes [12], while enhancing bleomycin A 2 cytotoxicity and DNA strand breakage in murine leukemia cells and human head and neck carcinoma cells [13] and imparing corneal wound healing [14].…”

“…Other local anesthetics appear to have similar effects on various cell types [9][10][11][12][13][14]. Prilocaine was found to induce apoptosis in mouse and human osteoblastic cells [11] and lidocaine was shown to inhibit the proliferation of normal human fibroblasts, endothelial cells and keratinocytes [12], while enhancing bleomycin A 2 cytotoxicity and DNA strand breakage in murine leukemia cells and human head and neck carcinoma cells [13] and imparing corneal wound healing [14]. Dibucaine was shown to induce apoptosis in a neuroblastoma cell line [9] and in HL-60 [10] cells through activation of various caspases in conjunction with the release of cytochrome c from mitochondria induced by a processed product of Bid and depolarization of the mitochondrial membrane potential [10].…”

“…There are relatively fewer in vitro studies investigating the effects of local anesthetics on cell proliferation, apoptosis and gene expression [9][10][11][12][13][14]. It was reported that ropivacaine was shown to inhibit the proliferation of nontransformed cultured adult human fibroblasts, endothelial cells, keratinocytes [12] and primary rat fibroblast [6], as well as that of the transformed monkey kidney Vero cells [6] and human colon cell lines [15].…”

Lidocaine-induced apoptosis and necrosis in U937 cells depending on its dosage

“…However, it is also known that they have other beneficial or adverse effects on a variety of cellular activities such as wound-healing, thrombosis, inflammatory responses, and cellular toxicity (4,5,16). One of the cellular toxicities of local anesthetics, an adverse effect, is an induction of necrosis or apoptosis in non-neuronal as well as neuronal cells in vivo and in vitro (4,10,12,22,27). It has been reported that administration of local anesthetics in vivo induced cell death in muscle cells (10) and neuronal cells (12).…”

<i>In vitro</i> oral epithelium cytotoxicity and <i>in vivo</i> inflammatory inducing effects of anesthetic rice gel

“…However, many studies have shown their cytotoxicity towards several cultured cells (13,14). Cytotoxicity of local anesthetic including lidocaine and prilocaine has been reported on mesenchymal stem cells and osteoblastic cells as well as human oral and tumor cells (15)(16)(17)(18). Even though these local anesthetics have been long used in several complications, there is inadequate information about cytotoxic activity of these agents particularly for dental applications that the use of local anesthetics has been increased.…”