## 2 Pharmacology of Piperine: In Vitro and In Vivo Studies
### Anti-Inflammatory Effect  
Piperine decreases liver marker enzymes activity (aspartate transaminase (AST), alanine, transaminase (ALT), and alkaline phosphatase (ALP)) in acetaminophen-challenged mice, indicating its hepatoprotective and antioxidant effects [24]. Piperine decreases blood urea nitrogen (BUN), creatinine, and malondialdehydedehyde (MDA), and increases superoxide dismutase (SOD), glutathione peroxidase (GPx) in the kidney of lead acetate-treated nephrotoxic rats [25]. Similar results were observed in cadmium-induced oxidative stress in cultured human peripheral blood lymphocytes in a HFD [26] and antithyroid drug-induced hyperlipidemic rats [27]. These enzymatic studies also revealed the anti-inflammatory activity of piperine. The administration of piperine to rats before irradiation significantly abolished the radiation-induced alleviation in lungs catalase (CAT) and GPx activities, reduced GSH content and significantly limited the elevation in serum tumor necrosis factor-\(\alpha\) (TNF-\(\alpha\)), interleukin-1\(\beta\) (IL-16) and interleukin-6 (IL-6) levels, which demonstrated its protective function from x-rays [28]. Piperine inhibits LPS-induced expression interferon regulatory factor (IRF)-1 and IRF-7 mRNA, phosphorylation of IRF-3, type 1 IFN mRNA, and reduces the activation of signal transducer and activator of transcription (STAT)-1. The results indicate that piperine is a potential molecule for treating lipopolysaccharide-induced inflammation [29]. Similarly, [PERSON] et al. [30] reportedthat piperine inhibits LPS-induced TNF-\(\alpha\), IL-6, IL-1\(\beta\), and prostaglandin E2 (PGE-2) production in BV2 microglial cells. In the human peripheral blood mononuclear cells (PBMCs), piperine was found to inhibit IL-2 and interferon gamma (IFN-x) [31]. Piperine inhibits the production of PGE2 and NO induced by LPS while decreasing TNF-\(\alpha\), inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2) in RAW264.7 cells, resulting in anti-inflammatory activity [32]. In the model of LPS-induced inflammation of nucleus pulposus cells, piperine inhibited IL-1\(\beta\), TNF-\(\alpha\), IL-6, expression of iNOS, activities of matrix metalloproteinases MMP-3 and MMP-13, ADAMTS-4 mRNA, and ADAMTS-5 mRNA [33].  
In an arthritis animal model, at doses of 10 and 100 \(\upmu\)g/mL, piperine inhibits the IL-6, MMP-13, activator protein 1 (AP-1) and reduces PGE2 in a dose-dependent manner, and significantly reduced nociceptive and arthritic symptoms in piperine-treated rats. Histological investigation revealed reduced inflammatory area in the ankle joints [34]. Piperine downregulates IL-1, MMP-8, and MMP-13 in periodontitis, leading to protective effects on inflammation, alveolar bone loss, bone microstructures, and collagen fiber degradation in experimental periodontitis [35]. [PERSON] et al. [36] reported that piperine suppresses cytosolic phospholipase A2 (cPLA2) and inhibits thromboxane A2(TXA2) synthase, but not of COX-1, in collagen-stimulated platelets. It also inhibits the lipopolysaccharide-induced generation of PGE2 and PGD2 in RAW264.7 cells by suppressing the activity of COX-2, without effect on cPLA2 [36]. Piperine reduces proinflammatory cytokines IL-1\(\beta\), IL-6 and TNF-\(\alpha\), COX-2, nitric oxide synthase (NOS-2), and nuclear factor kappa B (NF-\(\upkappa\)B) in the cerebral ischemia-reperfusion-induced inflammation rat model [34].  
In one study, performed on rats, piperine was found to reduces SOD, CAT, GPx, and GR, and increases hydrogen peroxide generation and lipid peroxidation in the epididymis, thus having a negative effect to redox state and affecting fertility [37].