It was found and analyzed that both pipecuronium and vecuronium could inhibit the expression of cd11b on neutrophil surface when the concentration of pipecuronium and vecuronium were close to the clinical maintenance concentration (low concentration of the test group). The expression of cd11b was not affected by the concentration of pipecuronium and vecuronium. Lipopolysaccharide could significantly activate the expression of cd11b on neutrophil surface. Vecuronium and pipecuronium did not affect the expression of cd11b induced by lipopolysaccharide activation on neutrophils. The whole blood sample was used and the incubation temperature was kept at 37℃, which was close to the physiological internal environment of the organism as far as possible, so as to avoid the damage of the physiological internal environment caused by the pure separation of neutrophils, which resulted in the decrease of the activity of the isolated neutrophils. The whole blood pattern can effectively maintain the physiological activity of neutrophils. It provides a good reference for further clinical trials in vivo. However, all kinds of immune cells and various immune factors in whole blood may be involved in regulating the expression of cd11b, and the whole blood environment may also affect the correctness of the conclusions. Krouholz w et al studied the effect of pancuronium bromide on the adhesion of neutrophils through in vitro experiments. They were 1.3µg•ml− 1 and 0.5µg•ml− 1 respectively. It was found that both clinical concentrations of pancuronium could directly affect neutrophil adhesion to neutrophil inhibition and significantly affect the chemotaxis of neutrophils to inflammatory sites. The inhibitory effect of high concentration of pancuronium was stronger. However, low concentration of vecuronium and pipecuronium could effectively inhibit the expression of adhesion molecule cd11b on neutrophil surface, but the high concentration of drug had no effect on the expression of adhesion molecule on neutrophil surface. This is different from the conclusion of krumholz w et al. It is speculated that the effect of different drugs on neutrophil function is not concentration dependent, or there are many other influencing factors or mechanisms. In this study, three kinds of muscle relaxants used high and low concentrations, close to the clinical induction concentration and muscle relaxant maintenance concentration respectively, as to whether the other concentrations of drugs can reach statistically significant conclusions need to be further experimental proof. In this paper, the random block design of the same blood sample is used to reduce the test error effectively. Due to the use of small sample data in the experiment, the exact conclusion remains to be demonstrated by repeated tests of multiple concentrations in large samples.
Known some anesthetics have certain inhibition of neutrophil respiratory burst, Muhling J et al found midazolam can change neutrophils in the free amino acids and alpha keto acid content, and significantly reduce the immune function index superoxide anion (O2) the formation of hydrogen peroxide (H2O2), and release MOP (myeloperoxidase, myeloperoxidase), midazolam dose dependent decrease peripheral blood neutrophils macrophages breathing outbreak. Drugs can directly or indirectly affect intracellular Ca2 + regulation of cellular function, and Nishina K et al found that intravenous anesthetics may reduce neutrophil function by reducing intracellular Ca2 + concentration. The mechanism of action may be that intravenous anesthetics, by virtue of their high lipid solubility, accumulate on the cell membrane, change the physical structure of the membrane, and cause the membrane enzyme action, especially the inhibition of protein kinase C, to change the cell function. In this study, two kinds of steroidal muscle relaxants, pipecuronium and vecuronium, were observed to slightly increase the expression of cell surface adhesion molecules, suggesting that steroidal muscle relaxants may change the cell membrane homeostasis, which is similar to the mechanism of action of intravenous anesthetics on the cell membrane.
In this study, only two steroidal muscle relaxants with low concentration showed inhibitory effect on CD11b expression, so it was speculated that the anti-inflammatory effect of steroidal muscle relaxants might be related to the presence of androstane structural groups. Krumholz W et al found that pancuronium bromide had an inhibitory effect on the adhesion of neutrophils in vitro, and the present study also found that vecuronium bromide and pipecuronium bromide with androstane structural groups could inhibit the expression of adhesion molecules, suggesting that the anti-inflammatory effect of muscle relaxants is related to its androstane structural groups.
As a water-soluble glycosylated lipid complex, lipopolysaccharide is the main component of the cell wall of gram-negative bacteria. LPS is an important mononuclear macrophage and neutrophil strong activator, which can cause the high expression of CD11b on the neutrophil surface and lead to the formation and activation of inflammatory responses by TNF-α、IL-1、IL-6、IL-8 and NO, and other pro-inflammatory factors. This experiment adopts the lipopolysaccharide as activators of whole blood in vitro, and whole blood 37℃ in lipopolysaccharide common incubation after 2 hour, by observing and analyzing the CD11b expression of complete blood found that CD11b expression significantly higher in the surface of neutrophils, the mechanism may be related inflammatory response is a combination of a variety of channels, speculate that the main way to have: (1) LPS activated the Nuclear Factor Kappa B, launch a variety of inflammatory related gene transcription, inflammatory factor to further activate the surface of the PMN CD11b/CD18 expression. CD11b/CD18, through transmembrane signal transduction, promotes the activity of inflammatory transcription factor -- Nuclear Factor Kappa B, inhibits cell apoptosis, and further promotes the increase of inflammatory gene expression products. These inflammatory mediators further up-regulate the expression of CD11b/CD18. LPS activated PMN, IκBα(nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, alpha) first significantly degraded, and then synthesis increased, inhibiting the activation effect of Nuclear Factor Kappa B, which led to a sharp increase in the initial expression of CD11b/CD18, followed by a decrease in the expression rate. (4) this test used whole blood environment, lipopolysaccharide in addition to the activation of neutrophils may also effect on macrophages and other mononuclear cells induced by producing a variety of inflammatory cell factor mutual activation of signaling pathways, before cascade amplification effect and activate CD11b expression. Induced by lipopolysaccharide CD11b express function strong, two kinds of steroid used in this test and muscle relaxants anti-inflammatory effects weak cannot restrain the effect of the activation. CD11b expression on lymphocyte surface was also observed to be stable, indicating that lipopolysaccharide did not affect lymphocyte activation, which may be related to the specificity of transcription factor-Nuclear factor-kappaB(NF-kB) or negative transcriptional feedback regulation of I B in lymphocyte, as well as other possible intermediate mechanisms.
In this study, it was found that two steroidal muscle relaxants could inhibit the expression of adhesion molecules at clinical microdose, which was consistent with the results of Krumholz W et al. 's study on pancuronium bromide.The results showed that vecuronium bromide and pipecuronium bromide had only a slight inhibitory effect on the expression of adhesion molecules on the surface of neutrophils, but could not effectively affect the activation and expression of lipopolysaccharide, and had a concentration-dependent effect.The muscle relaxants vecuronium bromide, pancuronium bromide and pipecuronium bromide have similar structures with acetyl groups, and choline receptors interact with the postsynaptic membrane by means of them.Pancuronium bromide has two functional groups.As a derivative of pancuronium bromide, vecuronium only lacks n-methyl groups at 2 positions. This structure leads to the easy hydrolysis reaction of the acetic acid root at C3 position, and the stability of the solution is lower than that of pancuronium bromide, and the action time is shorter.The structural difference between piperazine ring and pancuronium is that piperazine ring exists in the former and piperazine ring in the latter.Obviously, except for a few secondary groups, the molecular structure of the three steroidal muscle relaxants is very similar, and all of them can show some anti-inflammatory effects.
The slight inhibitory effect of steroidal muscle relaxants on neutrophil function was speculated to be related to the following mechanism by combining the results of various drugs on neutrophil adhesion. The structure of myasthenic steroids contains androstane like structural groups similar to steroid hormones, with mild anti-inflammatory and immunosuppressive functions.[9.10] The steroids muscle relaxants may change the physical structure of the cell membrane, causing the membrane enzyme action, especially the inhibition of protein kinase C, leading to the opening and closing of the ion channel, changing the PH value of intracellular acid and base, leading to changes in cell function.[8, 11, 12, 13] Steroidal muscle relaxants can reduce integrin expression by inhibiting intracellular receptor signaling pathway and kinase phosphorylation.[14, 15] In addition, nitric oxide synthase is regulated to inhibit the production of nitric oxide. It shall directly or indirectly inhibit the generation of transcription factors (nf-kb) and transcription factor activated protein-1 (ap-1).[10, 17]
In addition, the whole blood contains a small amount of lactoferritin (LF), which is an important immune regulator with anti-inflammatory and antioxidant properties and can reduce the release of free radicals.Neutrophil mast cells contain large amounts of LF receptors on their surface, and LF binds to receptors on the cell surface to regulate its immune function.Studies have found that LF can be expressed in mast cells and inhibit the IgE induced mast cell degranulation reaction, stabilize the inhibition of trypsin, and inhibit mast cell activation during allergy. Steroid muscle relaxants may, like glucocorticoid structures, up-regulate the expression of steroid receptor target genes and the transcription of lactoferritin, thereby inhibiting the expression of CD11b on the surface of neutrophils.
In conclusion, a certain dose of vecuronium and pipecuronium may inhibit the expression of CD11b on neutrophil surface. However, the effect of high concentration of vecuronium bromide and pipecuronium bromide in this experiment was not obvious, and the concentration and test method of the drug may be closely related to its results, and the specific mechanism still needs to be studied in a large sample.