exposure can show a wide range of results. The main results of phorbol exposure are tumor promotion and inflammatory response. Although phorbol is not a carcinogen itself, it greatly enhances the action of other substances and promotes tumor proliferation. PKC is a key component in biological pathways controlling cell growth and differentiation. When phorbol esters bind to PKC, cell proliferation pathways are activated. This effect greatly promotes tumors when the cells are exposed to even a sub-carcinogenic amount of a substance. PKC is also involved in activation of inflammation pathways such as the NF-κB pathway. Thus, exposure to phorbol products can induce an inflammatory response in tissues. Symptoms can include edema and pain, especially to the skin and mucus membranes. While phorbol itself does not have irritant activity, nearly all phorbol esters are highly irritant, with a wide range of half-maximal inhibitory concentration (IC50) values. The median lethal dose (LD50) of phorbol esters for male mice was found to be about 27 mg/kg, with the mice showing hemorrhage and congestion of pulmonary blood vessels, as well as lesions throughout the body. Total synthesis A total synthesis of enantiopure phorbol was developed in 2015. While this synthesis will not replace natural isolation products, it will enable researchers to create phorbol analogs for use in research, especially creating phorbol derivatives that can be evaluated for anti-cancer activity. Previously, the difficulty with synthesizing phorbol had been creating C–C bonds, especially in the six-membered ring at the top of the molecule. This synthesis starts from (+)-3-carene, and uses a series of 19 steps to eventually create (+)-phorbol. Uses in biomedical research and treatments Because of their mechanism of action, phorbol esters can be used to study tumor proliferation and pain response. TPA is most commonly used in the laboratory to induce a cellular response. For example, TPA can be used to measure response to pain and test compounds that may mitigate the inflammatory response. TPA and other phorbol esters can also be used to induce tumor formation and elucidate its mechanism. Some phorbol esters such as tigilanol tiglate have shown anti-cancer activities and can potentially be used as a defense against certain viruses and bacteria. TPA, together with ionomycin, can also be used to stimulate T-cell activation, proliferation, and cytokine production, and is used in protocols for intracellular staining of these cytokines. Phorbol, in the form of croton oil, is also in folk medicine