What Is a Kallikrein?

That is, kallikrein (KLK), which was discovered by Kraut and others in the pancreas in 1930, was named Kallikrein. In the past 30 years, with the development and application of molecular biology and cell biology technology, the kallikrein kinin system (KKS) has been discovered as a complex endogenous multi-enzyme system that participates in the regulation of cardiovascular , Kidney, nervous system and other physiological functions are closely related to the occurrence of heart disease, kidney disease, inflammatory response, cancer and other diseases.

Kallikrein

which is

First reported by Abelous et al. [1] in 1909

KKS is one of the main blood pressure systems in the body.

KLK in plasma

A large number of studies have shown that KKS plays a very important role in the pathogenesis of various diseases of the cardiovascular system, such as hypertension, heart failure and myocardial ischemia, LVH and endothelial dysfunction. With the deepening of people's understanding of KKS, not only in cardiovascular, but also in other pathological processes, it has gradually become a research hotspot. Specific receptors are becoming new targets for research, and the advent of corresponding antagonists will become a new generation of new and more selective drugs for the treatment of cardiovascular disease, inflammation, pain and immune diseases.

references

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Biological properties and mechanism of tissue kallikrein

Kallikrein in humans includes plasma kallikrein and tissue kallikrein, both of which are converted from prekalikrein and prokallikrein, respectively. Plasma kallikrein catalyzes the hydrolysis of high molecular kininogen to produce bradykinin and kallidin. In humans, tissue kallikrein is also known as pancreatic / kidney kallikrein [4], which can catalyze the hydrolysis of low-molecular kallikrein to produce pancreatic kallikrein. The carboxyl terminus of bradykinin and pancreatic kallikrein hydrolyzes Arg under the action of kallikrein I to generate des-Arg_-BK and des-Arg_-kallidin, respectively. The latter is still biologically active and requires angiotensin converting enzyme or amino group. Peptidase can be completely inactivated, and kallikrein mainly binds to B protein R and B protein R which play a role in G protein. B2 R is a housekeeping gene expression and is the main receptor for kallikrein under normal conditions. It is sensitive to bradykinin and pancreatic kinin. B1R is induced under inflammation and ischemia, and is responsible for des-Arg-BK is sensitive, in which B1 R is more sensitive to des-Arg_-kallidin than des-Arg-BK. It is thought that B1R may be involved in the inflammatory response and improvement of circulation at the site of injury, and play an important role in neovascularization. After kinin binds to the receptor, it activates the NO-CGMP and PG-CAMP pathways, thereby regulating the release of biologically active substances such as NO and PG to participate in the regulation of multiple organ functions and multiple disease processes, such as inhibition of apoptosis, inflammation, and hypertrophy Fibrosis promotes the generation of new blood vessels in the heart, kidney, and cerebral blood vessels and the generation of new nerves in the brain.

Protective effects of tissue kallikrein on cardiovascular and kidney

Human tissue kallikrein (HTK) is widely present in human kidney, cardiovascular, central nervous system, pancreas, intestine and other organs, and its metabolites bind to receptors to exert its extensive pathophysiology. Among them, HTK has the most research on cardiovascular and renal diseases.

The kallikreinkinin system (KKS) plays an important role in maintaining normal blood pressure and protecting the heart. Defects in the kallikreinkinin system (KKS) can cause hypertension. Berry in 1989 A family study has shown that human urinary kallikrein (HUK) reduces the risk of hypertension. Many animal experiments of hypertension or myocardial ischemia-reperfusion (I R) models have shown that adenovirus-based human tissue kallikrein gene (ad.htk) transduction can reduce hypertension and relieve myocardial hypertrophy and fibers It can also improve cardiac function, reduce the scope of myocardial infarction, and reduce ventricular fibrillation and apoptosis after myocardial I R.

Human urinary kallikrein (HUK) is a significant renal vasodilator, diuretic, and sodium excretory agent that protects the kidneys. Decreased HUMAN URINARY KALLIKREIN (HUK) can cause mild nephropathy in hospitalized patients, severe cases can cause severe renal failure, and the kallikreinkinin system (KKS) can inhibit inflammation and oxidation Enzymes to fight kidney failure caused by high salt diets or drugs.

Protective effect of tissue kallikrein on brain tissue

In humans, tissue kallikrein has been shown to be distributed on thalamus, hypothalamus, brain gray matter, neurons of the brainstem reticulum and adenoid pituitary cells and choroid plexus cells. B2R is expressed on neurons in human astrocytes, oligodendrocytes, microglia, cerebrovascular endothelial cells, cerebral cortex, striatum, thalamus, and hypothalamus. B1R is expressed in neurons and basal arteries of the thalamus and hypothalamus. In vitro studies have shown that human B1R is present in vascular endothelial cells, aorta smooth muscle cells, coronary arteries and muscular arterioles. B1R expression is up-regulated during ischemia and other injuries or inflammation. These all provide the premise for tissue kallikrein to protect brain tissue through the metabolites kallikrein and B1R and B2R. The specific neuroprotective effect and its mechanism of action are as follows:

1 Dilate cerebral arteries, improve blood and oxygen supply to ischemic brain tissue

The pathophysiology of cerebral ischemia has been studied in depth, and various theories have been proposed, but so far no mechanism can fully elucidate the damage mechanism of cerebral ischemia. It is believed that the molecular mechanisms involved in cerebral ischemic injury include the release of excitatory amino acids, the homeostasis of calcium ions, the formation of free radicals, the activation of proteases, and the mediation of NO.

The role of NO in cerebral ischemic damage has been a research hotspot. NO has dual effects of neuroprotection and neurotoxin. It is thought that the dual role of NO is related to its source. NO is catalyzed by L-arginine synthesis by NOS. NOS can be divided into structural (cNOS) and inducible (iNOS). CNOS includes endothelial (eNOS) and neuron (nNOS). Experiments have shown that the NO formed from iNOS and nNOS overexpression is neurotoxic, while the NO produced from eNOS has neuroprotective effect. So drugs that can increase NO by up-regulating eNOS can have neuroprotective effects.

B1R and B2R are special regulatable G protein-coupled receptors, and they have been confirmed to have the same cellular signal transduction pathway in endothelial cells. When kinin binds to B1 R or B2 R, the G protein coupled to the intracellular end of the receptor activates phosphatase C (PLC), and the PLC further hydrolyzes inositol 4,5-bisphosphate (IP3). The cytoplasm binds to the IP3 receptor in the sarcoplasmic reticulum, causing Ca2 + release in the reservoir, extracellular Ca2 + influx, increasing intracellular Ca2 +, and finally activating eNOS to produce NO. Increased intracellular Ca2 + also activates phospholipase A2 (PLA2), which induces PGI2. Lamontagne, Be & acute; lichard, etc. pointed out that des-Arg_-BK vasodilator effect is at least partially mediated by NO, and PG does not seem to be important. The role of kinin in dilating the cerebral arteries comes in part from the release of NO.

Patients with acute stroke have increased pancreatic kinin in the peripheral circulation during the 8 days after the onset of the ischemic attack. Studies by Simone and others showed that 22 patients with middle cerebral artery occlusion with larger infarcts had higher tissue kallikrein concentrations and higher pancreatic kallikrein than 14 normal adults. These all indicate that tissues of kallikrein and pancreatic kallikin are activated in ischemic brain tissue. Pancreatic kinin has a significant vasodilator effect. Pancreatic kinin and its metabolite des-Arg_-kallidin can bind to B2R and B1R, respectively, and release NO to expand cerebral arteries. In normal humans and animals, vasodilator action is mainly mediated by B2R; in the case of injury such as inflammation or ischemia, vasodilator action is mainly mediated by newly expressed B1R. As in pathological conditions, B1R shows a more pronounced coronary dilation effect than B2R.

During ischemic injury such as acute cerebral infarction, vascular cells in the ischemic area are induced to generate B1R. At this time, kinin is combined with B1R to expand arterial blood vessels in the brain tissue of the ischemic area, thereby improving the blood supply and oxygen supply of ischemic brain tissue. .

2 Promote the generation of new blood vessels in ischemic brain tissue

In patients and animal models of peripheral vascular disease, Kallikrein-kinin system (KKS) was up-regulated. Kallikrein-kinin system (KKS) was used in myocardial / limb ischemic diseases. It plays an important role in promoting neovascularization and inhibiting apoptosis. There is a theory that kinin has a long-term protective effect on ischemic tissues by enhancing angiogenesis. Local transduction of HTK gene can cause angiogenesis and promote tissue recovery in this area. In vivo experiments show that HTK gene transduction can promote corneal neovascularization and capillary proliferation in rabbits. Studies have shown that low-dose (106 PFU) ad.htk transduction into mice can promote the growth of muscle capillaries and arteries in limbs, and 107 PFU ad.htk can further expand the microvasculature. In diabetic mice induced by streptozotocin, topical administration of KLK can stop the process of microvascular reduction in hindlimb skeletal muscle. This effect is achieved by inhibiting apoptosis and promoting vascular regeneration. After inhibiting KLK with tissue kallikrein inhibitor KLK-binding protein, it was observed that it inhibited the increase of capillary endothelium and induced apoptosis, and finally inhibited the formation of new blood vessels.

In vitro studies have found that kinin activates endothelial nitric oxide synthase (eNOS) via the IP3-AKt / protein kinase B (ie, IP3-AKt-B) pathway, thereby allowing vascular endothelial growth factor receptors to pass eNOS The mediation causes the formation of stroma endothelial cells. In vivo studies have shown that AKt-B and eNOS are functionally related to the neovascularization pathway induced by ad.htk. Ad.htk-induced kallikrein and vascular endothelial growth factor A work together to induce angiogenesis, produce NO, and relax blood vessels.

Pharmacological studies have shown that B1R plays a role in capillary proliferation. B1R not only mediates the growth and survival of endothelial cells in ischemic injury (kinin can effectively attract leukocytes, which are required for the production of endothelial cell growth factors), but can also participate in ischemia by increasing the exudation of extravascular plasma proteins Post-neovascularization (these proteins provide a temporary scaffold for angiogenesis). Recent studies have shown that hereditary B1R deficiency cannot repair new blood vessels. Therefore, B1R plays an important role in promoting the generation of neovascularization in ischemic tissue.

During acute cerebral ischemia and cell injury, the expression of B1R in ischemic cells is up-regulated, tissue kallikrein binds to B1R through the metabolite des-Arg_-kallidin, and further activates the endothelium through the IP3-AKt-B or calmodulin pathway Nitric oxide synthase (eNOS), thereby promoting the formation of new blood vessels in ischemic brain tissue.

3 Promote glial cell migration and inhibit apoptosis, reduce inflammatory cell invasion

Different animal models have shown that tissue kallikrein (KLK) reduces heart and kidney brain organ damage by inhibiting apoptosis and inflammatory cell invasion. In the model of cerebral ischemia caused by blocking middle cerebral artery (MCAO) in rats, Julie Chao and Lee Chao can significantly reduce ischemia-induced neurological damage, reduce the area of cerebral infarction, and promote Glial cells survive and migrate to the ischemic penumbra area and center, reduce the apoptosis of nerve cells and glial cells, infiltrate inflammatory cells, promote neovascularization and nerve cell regeneration, thereby improving survival rates. Continuous cerebral micropump infusion of human tissue kallikrein after MCAO in the brain has a direct effect on the recovery of neurological functions such as movement disorders caused by ischemia-reperfusion.

MCAO-induced cerebral ischemia-reperfusion damages the blood-brain barrier, and KLK genes or proteins can enter the ischemic injury area through the blood-brain barrier, thereby exerting neuroprotective effects. Morphological analysis showed that KLK gene transduction enhanced survival and promoted glial cell migration to the ischemic penumbra area and center. After the introduction of KLK gene, the cell survival rate increased and increased the levels of NO and phospho-Akt, Bcl-2 in the brain, reduced caspase-3 activation, reduced NAD (P) H oxidase activity, and inhibited superoxide Be relevant. This indicates that the protective effect of KLK gene or protein transfer on cerebral ischemic injury does not depend on its vasodilator and blood pressure lowering effects, but through the following ways: to promote the survival and migration of glial cells, and to inhibit oxidative stress and inhibition Akt-Bcl-2 signaling pathway to inhibit apoptosis.

Studies show that kallikrein's effects on cell migration and apoptosis can be blocked by the B2 R antagonist itibante, suggesting that B2 R mediates these effects, and the role of B2 R in protecting ischemic stroke is also at the same time as B2 R deficiency Confirmed in rats. B2R-deficient mice show more infarct size and apoptosis, and more severe mobility disorders than wild-type mice after ischemia-reperfusion (I / R) injury. They had less white blood cell aggregation than wild type mice on day 1 of ischemia, but had more white blood cell aggregation than wild type mice on day 3 of ischemia. Studies have shown that early (brain ischemia 0.25 h to 6.25 h) use of B2R antagonists can reduce transient cerebral ischemic injury by inhibiting edema formation. The above shows that B2R has a dual effect: it promotes the invasion of inflammatory cells in the early stage of ischemia and causes increased vascular permeability, but in the later stage, it promotes Akt phosphorylation, reduces NAD (P) H oxidase activity, and inhibits superoxide to play a neuroprotective role .

4 Antagonizes vascular injury and inhibits arterial hypertrophy

Murakami et al. After transducing KLK gene locally in the left common carotid artery of rats after vascular balloon angioplasty, the intimal / medial ratio of the artery was significantly lower than that of the control group. It is NO dependent. In a mouse arterial remodeling model, Emanueli et al. Found that whole-body KLK genes reduce neointimal formation by altering vascular shear stress. Zhang et al., after transducing the KLK gene in a rat model of hypertensive cerebral hemorrhage caused by a high-salt diet, significantly relieved hypertension, inhibited arterial hypertrophy, and reduced cerebral hematoma. It is through these effects that tissue kallikrein releases hypertensive cerebral hemorrhage Played an important protective role in reducing animal mortality.