• 한국동물위생학회지
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• 제41권3호
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• pp.171-177
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• 2018

Purified bee venom was collected from colonies of honeybees (Apis mellifera L.) using a bee venom collector under sterile conditions and then purified under strict laboratory conditions. Purified bee venom contained $63.9{\pm}5.4%$ melittin, $10.9{\pm}1.6%$ phospholipase A2, and $2.3{\pm}0.3%$ apamin. Purified bee venom has various anti-bacterial, anti-inflammatory and immunostimulating effects. In this study, we evaluated purified bee venom which are mammary gland cells, MAC-T cells are used to increase the synthesis of milk protein. Purified bee venom promoted the proliferation of MAC-T cells at concentrations below $1{\mu}g/mL$, but cytotoxicity at $10{\mu}g/mL$ and above. As a result of the increase in the synthesis of ${\beta}-casein$, a milk protein after treatment with MAC-T cells at a concentration of the bee venom without cytotoxicity, the ${\beta}-casein$ content in the cell culture was increased when treated at a concentration of 1 ng/mL or more. In addition, it was confirmed that purified bee venom significantly increased the expression of bovine ${\beta}-casein$ (bCSNB) mRNA, a ${\beta}-casein$ synthesis gene, at a concentration of 1 ng/mL or more. These results suggest that purified bee venom can be used to increase the production of livestock by ultimately increasing the expression of milk protein.

• The Korean Journal of Physiology and Pharmacology
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• 제2권2호
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• pp.225-232
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• 1998

To investigate the possible involvement of outward potassium ($K^+$) currents in nitric oxide-induced relaxation in intestinal smooth muscle, we used whole-cell patch clamp technique in freshly dispersed guinea-pig ileum longitudinal smooth muscle cells. When cells were held at -60 mV and depolarized from -40 mV to -50 mV in 10 mV increments, sustained outward $K^+$ currents were evoked. The outward $K^+$ currents were markedly increased by the addition of 10 ${\mu}M$ sodium nitroprusside (SNP). 10 ${\mu}M$ S-nitroso-N-acetylpenicillamine (SNAP) and 1 mM 8-Bromo-cyclic GMP (8-Br-cGMP) also showed a similar effect to that of SNP. 1 mM tetraethylammonium (TEA) significantly reduced depolarization-activated outward $K^+$ currents. SNP-enhanced outward $K^+$ currents were blocked by the application of TEA. High EGTA containing pipette solution (10 mM) reduced the control currents and also inhibited the SNP-enhanced outward $K^+$ currents. 5 mM 4-aminopyridine (4-AP) significantly reduced the control currents but showed no effect on SNP-enhanced outward $K^+$ currents. 0.3 ${\mu}M$ apamin and 10 ${\mu}M$ glibenclamide showed no effect on SNP-enhanced outward $K^+$ currents. 10 ${\mu}M$ 1H-[1,2,4]oxadiazolo [4,3-a]quinoxaline-1-one (ODQ), a specific inhibitor of soluble guanylate cyclase, significantly blocked SNP-enhanced $K^+$ currents. We conclude that NO donors activate the $Ca^{2+}-activated$ $K^+$ channels in guinea-pig ileal smooth muscle via activation of guanylate cyclase.

• 동의생리병리학회지
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• 제27권1호
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• pp.92-98
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• 2013

Bee Venom(below BV) has been used in alternative medicine to treat the diseases, such as pain diseases. BV contains a variety of peptides, including melittin, apamin, adolapin, MCD peptide, enzymes(i.e. PLA2), amines(i.e. histamine and epinephrine), and nonpeptide components. The two main components of BV are melittin and PLA2. The cell cytotoxic effects through the activation of PLA2 by melittin have been suggested to be the critical mechanism for the depress of cancer cell. Melittin and PLA2 have been reported to induce apoptosis and to possess anti-cancer effects and neurite outgrowth in PC12 cells. Analysis of proliferation was confirmed by MTT assay. BV decreased cell number through dose- and duration-dependent manner and these effects are apparent at a concentration of 3 ${\mu}g/ml$. To observe which signaling molecules will be activated by BV, phosphorylation of ERK, p38 MAPK, JNK and ERM were examined by Western blot analysis. To study the long term effect of BV in human gastric adenocarcinoma cell lines, the image of cells treated with BV for 4 days were obtained. BV was shown to exhibit anti-cancer activity in human gastric adenocarcinoma cell lines at a broad range of concentrations of 3 ${\mu}g/ml$. ERK, p38 MAPK and JNK were found to increase in BV treated cells. However, ERM which known to be involved in the cell death, was gradually decreased to 30minutes after addition 3 ${\mu}g/ml$ of BV. These results provide a possible BV-induced inhibitory signal for cancer proliferation that is initiated by the decrease in ERM activity. Moreover, it is likely that the activation of ERK, p38 MAPK and JNK are required for the BV-induced inhibition of cancer proliferation.

• 대한한의학회지
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• 제21권3호
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• pp.3-8
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• 2000

1. Definition : Bee-venom therapy does not involve actual bee-stings: it is a treatment method with acquired bee-venom extract through the electric stimulus on the bee, It is injected subcutaneusly on the acupuncture point after refining, according to the diagnosis of constitution and disease. 2. History : Around 2000 B.C., records that Bee-venom was used for therapy were written in the medical book of Babylonia and Papirus of ancient Egypt. Hippocrates, who is called the father of Medicine, said that Bee-venom is Arcanum, which means mysteric medicine. In Oriental medicine, B.C. 200, there was a clinical record that the meat suspended in front of the bee house on the tree in order to get bee-venom, was attached on the lesion. 3. Mechanism of Action : There are two aspects: 1) The effect of stimulating acupunture point It is similar to the chemical moxa. I think that there are several methods of stimulating the acupuncture point： For example, a simple needle is a mechanical stimulus, Moxa is a heating stimulus and electric and Raser acupunture etc. And another stimulus: in the ancient orient, a chemical stimulus called Chungu(Tianjiu), is attached to the lesions by using grinded insects (ex. Mylaris phalerate PALL.) which have toxin. So Bee venom therapy is similar to this. 2) The effect of biochemical ingredients Bee venom consists of 40 kinds of ingredients. For example, me Iii tin, Apamin, Pospholipase A2, MCD peptide, Adolapin and so on. They have effects which have been proven through experimentation l) tonifying mechanism of the body through increasing hormon secretion 2) tonifying immune system through proliferation of WBC, lymphocytes, macrophage 3) anti-inflammatory reaction Therefore Bee venom therapy is the representative 3rd Medicine, which combined East & West medicine. 4. Application of disease : L.B.P and HIVD, O.A, R.A, degenerative arthritis, shoulder pain and other pain diseases. 5. Therapic methods : According to constitution and disease, proper concentration of bee venom is injected on acupunture point, 2 times a week. Generally one term is consisted of 15times. 6. Contraindication : Heart disease, TBc, DM, kidney disease(nephritis), pregnancy, woman in menstruation 3-4 persons per 100,000 persons may have severe allergic reaction.

• 한국응용약물학회:학술대회논문집
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• 한국응용약물학회 1994년도 춘계학술대회 and 제3회 신약개발 연구발표회
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• pp.288-288
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• 1994

1.목적 $K^{+}$ 통로 개방제인 cromakalim과 levcromakalim이 원형질막의 $K^{+}$ 통로를 개방시켜 막의 과분극을 일으킴으로서 강력한 혈관 이완작용을 일으킨다는 것은 주지하는 바다. 본 연구진들은 지난 2년간의 신약개발 연구사업을 통하여 cromakalim과 pinacidil 등 여러종의 K+통로개방제가 건전한 평활근 세포에서 phenylephrine뿐만 아니라 saponin처리에 의한 투과성 근세포 (permeabilized cells)에서 inositol 1,4,5-triphosphate (IP$_3$)에 의한 수축도 억제함을 보고 하였다. 본 연구에서는 이러한 수축작용에 대한 SPEX fluolog-2 spectrophotometer를 사용하여 돼지의 관상동맥 혈관근의 세포질내 $Ca^{++}$의 농도 ([$Ca^{++}$])의 변동을 관찰하였다. 정상 관상동맥 혈관근 조직에서 acetylcholine (0.1-1$\mu$M)에 의한 [Ca$^{++}$]농도의 증가와 b-escin 처리에 의한 skinned strip에서의 IP3 (1-5$\mu$M)에 의한 [Ca$^{2+}$]의 증가는 cromakalim과 levcromakalim의 전처치에 의하여 현저히 억제되었다. Skinned strip에서 이러한 K+ 통로 개방제에 의한 $IP_3$-요도 ($Ca^{2+}$)i 증가의 억제가 apamin (1-5 mM)과 glibenclamide (1$\mu$M)에 의하여 봉쇄되었으나, chrybdotoxin (0.1$\mu$M)에 의하여는 영향을 받지 아니하였다 한편 skinned strip에서 cromakalim은 GTP${\gamma}$s에 의한 [$Ca^{2+}$]i의 증가는 봉쇄하였으나 caffeine에 의한 [$Ca^{2+}$]i의 증가는 영향을 미치지 아니하였다. 이러한 연구결과로 보아 cromakalim과 levcromakalim과 같은 $K^{+}$ 통로 개방제가 세포막의 $K^{+}$ 통로를 개방하는 작용 뿐만 아니라 적어도 sarcoplasmic membrane에서 $Ca^{2+}$의 유리를 봉쇄함으로써 강력한 혈관 평활근 이완 작용을 나타내는 것으로 시사된다.

• The Korean Journal of Physiology
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• 제29권2호
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• pp.233-241
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• 1995

The nonapeptide bradykinin has been shown to exhibit an array of biological activities including relaxation/contraction of various smooth muscles. In order to investigate the effects of bradykinin on the contractility and the electrical activity of antral circular muscle of guinea-pig stomach, the isometric contraction and membrane potential were recorded. Also, using standard patch clamp technique, the $Ca^{2+}-activated$ K currents were recorded to observe the change in cytosolic $Ca^{2+}$ concentration. $0.4 {\mu}M$ bradykinin induced a triphasic contractile response (transient contraction-transient relaxation-sustained contraction) and this response was unaffected by pretreatment with neural blockers (tetrodotoxin, atropine and guanethidine) or with apamin. Bradykinin induced hyperpolarization of resting membrane potential and enhanced the amplitude of slow waves and spike potentials. The enhancement of spike potentials was blocked by neural blockers. Both the bradykinin-induced contractions and changes in membrane potential were reversed by the selective $B_2$-receptor antagonist $(N{\alpha}-adamantaneacetyl-_{D}-Arg-[Hyp, Thy,_{D}-Phe]-bradykinin)$. In whole-cell patch clamp experiment, we held the membrane potential at -20 mV and spontaneous and transient changes of Ca-activated K currents were recorded. Bradykinin induced a large transient outward current, consistent with a calcium-releasing action of bradykinin front the intracellular calcium pool, because such change was blocked by pretreatment with caffeine. Bradykinin-induced contraction was also blocked by pretreatment with caffeine. From these results, it is suggested that bradykinin induces a calciumrelease and contraction through the $B_{2}$ receptor of guinea-pig gastric smooth muscle. Enhancement of slow wave activity is an indirect action of bradykinin through enteric nerve cells embedded in muscle strip.

• The Korean Journal of Physiology and Pharmacology
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• 제4권5호
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• pp.403-408
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• 2000

The outward currents elicited in hamster eggs by depolarizing pulses were studied. The currents appeared to comprise at least two components, a transient outward component $(I_{to})$ and a steady-state outward component $(I_{\infty}).\;I_{to}$ was transiently followed by the cessation of inward $Ca^{2+}$ current $(I_{Ca}),$ and its current-voltage (I-V) relation was a mirror image of that of $(I_{Ca}).$ Either blockade of $(I_{Ca})$ by $Co^{2+}$ or replacement of $Ca^{2+}$ with $Sr^{2+}$ abolished $I_{to}$ without change in $I_{\infty}.$ Intracellular EGTA (10 mM) inhibited $I_{to}$ but not $I_{\infty}.$ suggesting strongly that generation of $I_{to}$ requires intracellular $Ca^{2+}.$ Apamin (1 nM) abolished selectively $I_{to},$ indicatingthat $I_{to}$ is $Ca^{2+}-dependent\;K^+$ current. On the other hand, $I_{\infty}$ was $Ca^{2+}-independent.$ Both $I_{to}$ and $I_{\infty}$ were completely inhibited by internal $Cs^+$ and external TEA. The estimated reversal potential of $I_{to}$ was close to the theoretical $E_K.$ Taken together, both outward currents were carried by $K^+$ channels. From these results, $I_{to}$ is likely to be a current responsible for the hyperpolarizing responses seen in hamster eggs at fertilization.

• Journal of Acupuncture Research
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• 제20권4호
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• pp.53-65
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• 2003

Although the effect of Bee Venom has been reported, its mechanism has not been fully elucidated. Nitric Oxide(NO) is one of the free radicals and mediates in inflammation diseases. Chemokines contribute to the pathogenesis of several disorders such as allergic rhinitis and rheumatoid arthritis and so on. The objective of this study was to investigate the scavenging effect of Bee Venom on NO and on expression of chemokine genes. There was no significant NO scavenging effect in Crude Bee Venom, Apamin, Melittin, and MCD-peptide. The expression of chemokines was examined by RT-PCR using the human mast cell line(HMC-1), which is known to secrete and express chemokines. In order to investigate the protective effect of Bee Venom, HMC-1 cells were incubated with pretreatment of Bee Venom for 24 hrs and stimulated with 1 uM calcium ionophore A23178 for 2 hrs. RT-PCR analyses of chemokine genes showed that expressions of RANTES and MCP-1 were increased compared to the calcium ionophore-only treated group. But IL-8 and MCP-3 did not express increasing effect compared to control group. This study may provide important basic data on the possibility of the clinical treatment of Bee Venom in inflammation diseases.

• Advances in Traditional Medicine
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• 제2권2호
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• pp.87-93
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• 2002

The present study was aimed to examine the role of endothelium in the relaxant effect of hawthorn fruit extract of Crataegus pinnatifida in four different types of rat arteries, posterior cerebral communicating artery, right descending coronary artery, common carotid artery, and aorta. In $9,11-dideoxy-11{\alpha}$, $9{\alpha}-epoxy-methanoprostaglandin$ $F_{2{\alpha}}$ (U46619)-preconstricted arterial rings except for aorta, the extract produced endothelium-independent relaxations with similar potency. This relaxation was unaffected by pretreatment with $100\;{\mu}M\;N^G-nitro-L-arginine$ methylester (L-NAME, the nitric oxide synthase inhibitor), $3\;{\mu}M$ 1H-[l,2,4]oxadiazolo$[4,2-{\alpha}]$quinoxalin-1-one (ODQ, the guanylate cyclase inhibitor), or $10\;{\mu}M$ indomethacin (the cyclooxygenase inhibitor). Putative $K^+$ channel blockers (charybdotoxin plus apamin or glibenclamide) did not affect the extract-induced relaxation in cerebral or coronary artery rings. In contrast, in rat aortic rings the extract produced significantly smaller relaxant response in endothelium-denuded rings than that in endothelium-intact rings. Pretreatment with L-NAME or ODQ abolished the extractinduced endothelium-dependent aortic relaxation, whilst indomethacin $(3\;{\mu}M)$ had no effect. The present results indicate that hawthorn fruit extract possesses a vasorelaxing effect in cerebral, coronary and carotid arteries and this effect is independent of the presence of a functional endothelium. However, the extract-induced endothelium-dependent relaxation in rat aorta was mediated through endothelial nitric oxide and cyclic GMP-dependent mechanisms, suggesting that active components in the extract may act on endothelium to stimulate release of nitric oxide in large conduit arteries of the rats.

• The Korean Journal of Physiology and Pharmacology
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• 제13권5호
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• pp.361-365
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• 2009

Effects of quercetin, a kind of flavonoids, on the vasodilating actions were investigated. Among the mechanisms for quercetin-induced vasodilatation in rat aorta, the involvement with the $Ca^{2+}$ activated $K^+$ ($K_{Ca}$) channel was examined. Pretreatment with NE ($5\;{\mu}M$) or KCl (60 mM) was carried out and then, the modulation by quercetin of the constriction was examined using rat aorta ring strips (3 mm) at $36.5^{\circ}C$. Quercetin (0.1 to $100\;{\mu}M$) relaxed the NE-induced vasoconstrictions in a concentrationdependent manner. NO synthesis (NOS) inhibitor, NG-monomethyl-L-arginine acetate (L-NMMA), at $100\;{\mu}M$ reduced the quercetin ($100\;{\mu}M$)-induced vasodilatation from $97.8{\pm}3.7%$ (n=10) to $78.0{\pm}11.6%$ (n=5, p<0.05). Another NOS inhibitor, L-NG-nitro arginine methyl ester (L-NAME), at $10\;{\mu}M$ also had the similar effect. In the presence of both $100\;{\mu}M$ L-NMMA and $10\;{\mu}M$ indomethacin, the quercetin-induced vasodilatation was further attenuated by $100\;{\mu}M$ tetraethylammonium (TEA, a $K_{Ca}$ channel inhibitor). Also TEA decreased the quercetin-induced vasodilatation in endothelium-denuded rat aorta. Used other $K_{Ca}$ channel inhibitors, the quercetin-induced vasodilatation was attenuated by $0.3\;{\mu}M$ apamin (a SK channel inhibitor), but not by 30 nM charybdotoxin (a BK and IK channel inhibitor). Quercetin caused a concentration-dependent vasodilatation, due to the endotheliumdependent and -independent actions. Also quercetin contributes to the vasodilatation selectively with SK channel on smooth muscle.