plasma homocysteine (hcy) elisa measurement of blood from h-type hypertensive rats treated with huotan jiedu tongluo decoction

H-Tipi Hipertansif Sıçanlarda Huotan Jiedu Tongluo'nun Antihipertansif Etkileri

Hypertension, a major risk factor for heart attack, stroke, and renal failure, has become a significant public health challenge that affects 1 billion people worldwide1. Homocysteine (Hcy), a thiol group-containing amino acid, is a vital metabolic intermediary of methionine metabolism. Hypertension with elevated plasma Hcy levels is defined as H-type hypertension, which could be a significant risk factor for the occurrence and recurrence of cardiocerebrovascular diseases such as stroke2,3. Recent studies have reported that the co-residency of H-type hypertension could aggravate the side effects of cardiovascular and cerebrovascular diseases4. Notably, 75% of patients in China with H-type hypertension have primary hypertension, which seriously affects the quality of life5. At present, the treatment of H-type hypertension mainly includes Western medicine. However, it may cause certain adverse effects and poor compliance and can no longer meet the needs for the comprehensive management of H-type hypertension.
Traditional Chinese medicine (TCM) is a unique resource with a history of more than 2,000 years in China. Due to the unmet need for hypertension control in Western medicine, clinicians have begun to consider the potential role of TCM in the prevention and treatment of H-type hypertension6. Huotan Jiedu Tongluo Decoction (HTJDTLD) is a traditional Chinese medicine formula formulated by Professor Yue Deng, drawing from his extensive clinical expertise7. Over the course of more than 20 years of clinical application, HTJDTLD has demonstrated remarkable effectiveness in the treatment of cardiovascular and cerebrovascular diseases1. However, whether HTJDTLD has therapeutic effects in H-type hypertension has not been reported. Therefore, we aimed to explore the antihypertensive effects and specific mechanisms of HTJDTLD in rats with H-type hypertension and identify potential therapeutic drugs for the treatment of H-type hypertension.

Yazar Spotlight: Huotan Jiedu Tongluo Deoction'ı Antihipertansif Bir İlaç Olarak Keşfetmek

H-Tipi Hipertansiyonu Olan Sıçanlarda Huotan Jiedu Tongluo Dekaynatmasının Antihipertansif Etkileri ve Mekanizmaları

H-type hypertension has become a significant public health challenge of human. It mainly relies on Western medicine treatment, which can have sad effects. Our research explores the anti-hypertensive effects and mechanisms of Huotan Jiedu Tongluo decoction, a traditional Chinese medicine formula, in rats with H-type hypertension.Our study indicated that Huotan Jiedu Tongluo decoction had anti-hypertensive effects in rats with H-type hypertension. The underlying mechanism may involve inhibiting ER stress activation-induced apoptosis which provide an effective treatment option for patients. We will explode the potential biomarkers for early diagnosis and prognosis and develop novel therapeutic drugs for H-type hypertension in the future.

Huotan Jiedu Tongluo Dekotion ile Tedavi Edilen H-Tipi Hipertansif Sıçanlardan Alınan Kan Plazma Homosistein (Hcy) ELISA Ölçümü

Başlamak için, pıhtılaşmış kan içeren tüpleri 626 ila 1.409 G'de 20 dakika boyunca iki ila sekiz santigrat derecede santrifüjleyin. Sonra süpernatanı dikkatlice yeni bir tüpte toplayın. Süpernatanı eksi 80 santigrat derecede saklayın.Ardından, boş standart ve numune kuyuları ayarlayın. Seyreltilmiş standartlardan 50 mikrolitreyi ELISA plakasına pipetleyin. Daha sonra seyreltilmiş numunelerin 40 mikrolitresini numune kuyucuklarına aktarın.Numune kuyucuklarına 10 mikrolitre serum ekleyin. Şimdi, plakayı bir sızdırmazlık filmi ile kapatın. Kapalı plakayı 30 dakika boyunca 37 santigrat derecede bir inkübatöre yerleştirin.Tek mukavemetli bir yıkama çözeltisi elde etmek için 30 kez konsantre yıkama solüsyonunu damıtılmış su ile seyreltin. Ardından, sızdırmazlık filmini plakadan çıkarın ve sıvıyı atın. Şimdi, her kuyuyu beş kez 200 mikrolitre yıkama solüsyonu ile doldurun.Süpernatanı son kez attıktan sonra, plakayı kurulayın. 50 mikrolitre enzim reaktifini boş olanlar dışındaki tüm kuyucuklara pipetleyin. Ardından, plakayı 30 dakika boyunca 37 santigrat derecede bir inkübatöre yerleştirmeden önce bir sızdırmazlık filmi ile kapatın.Plakayı beş kez daha yıkadıktan sonra, renk geliştiricilerin her birinden 50 mikrolitre ekleyin, A ve B. İyice karıştırmak için plakayı hafifçe çalkalayın. Daha sonra plakayı düşük ışıkta 37 santigrat derecede 10 dakika inkübe edin. Şimdi, reaksiyonu sonlandırmak için her bir oyuğa 50 mikrolitre durdurma çözeltisi pipetleyin.Her bir kuyucuğun emilimini 450 nanometrede sırayla ölçün. Metiyonin grubundaki plazma homosistein konsantrasyonu, kontrol grubuna göre iki kat daha fazlaydı. Plazma homosistein konsantrasyonları tedavi gruplarında nispeten daha düşüktü.

plasma-homocysteine-hcy-elisa-measurement-blood-from-h-type

Research

JoVE Journal

Biochemistry

Plasma Homocysteine (Hcy) ELISA Measurement of Blood from H-Type Hypertensive Rats Treated with Huotan Jiedu Tongluo Decoction

90 Görüntüleme. Başlamak için, pıhtılaşmış kan içeren tüpleri 626 ila 1.409 G'de 20 dakika boyunca iki ila sekiz santigrat derecede santrifüjleyin. Sonra süpernatanı dikkatlice yeni bir tüpte toplayın. Süpernatanı eksi 80 santigrat derecede saklayın.Ardından, boş standart ve numune kuyuları ayarlayın. Seyreltilmiş standartlardan 50 mikrolitreyi ELISA plakasına pipetleyin. Daha sonra seyreltilmiş numunelerin 40 mikrolitresini numune kuyucuklarına aktarın.Numune kuyucukl...

Video: Huotan Jiedu Tongluo Dekotion ile Tedavi Edilen H-Tipi Hipertansif Sıçanlardan Alınan Kan Plazma Homosistein (Hcy) ELISA Ölçümü

The Antihypertensive Effects and Mechanisms of Huotan Jiedu Tongluo Decoction in Rats with H-Type Hypertension

H-type hypertension, which is a specific form of hypertension characterized by elevated plasma homocysteine (Hcy) levels, has become a major public health challenge worldwide. This study investigated the hypotensive effects and underlying mechanisms of Huotan Jiedu Tongluo decoction (HTJDTLD), a highly effective traditional Chinese medicine formula commonly used to treat vascular stenosis. Methionine was used to induce H-type hypertension in rats, and HTJDTLD was administered intragastrically. Then, the systolic and diastolic blood pressures of the caudal artery of rats were measured by noninvasive rat caudal manometry. Histological assessment of the aorta was performed by hematoxylin-eosin (HE) staining. Enzyme-linked immunosorbent assay (ELISA) was used to measure Hcy levels, and quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) and western blotting were used to determine the mRNA and protein levels of Glucose regulatory protein 78 (GRP78), Tumor necrosis factor (TNF) receptor-associated factor 2 (TRAF2), c-Jun N-terminal&#160;kinases (JNK), and caspase-3. The results showed that HTJDTLD significantly lowered blood pressure, alleviated histopathological lesions, and decreased Hcy levels after methionine treatment. Moreover, HTJDTLD significantly inhibited the gene and protein expression of GRP78, JNK, TRAF2, and caspase 3, which are involved mainly in the endoplasmic reticulum (ER) stress-induced apoptosis pathway. Overall, the results indicated that HTJDTLD had effective antihypertensive effects in rats with H-type hypertension and revealed the antihypertensive mechanisms associated with inhibition of ER stress-induced apoptosis pathway activation.

Here, we present a protocol to induce H-type hypertension and evaluate the antihypertensive effects of Huotan Jiedu Tongluo decoction (HTJDTLD) administered intragastrically. In rats with H-type hypertension, HTJDTLD had effective antihypertensive effects, possibly associated with inhibition of endoplasmic reticulum (ER) stress-induced apoptosis pathway activation.

H-type hypertension, which is a specific form of hypertension characterized by elevated plasma homocysteine (Hcy) levels, has become a major public health ...

Biyokimya

Treatment of Methionine-Induced H-Type Hypertensive Rats with Huotan Jiedu Tongluo Decoction (HTJDTLD)

To begin, segregate 50 hypertensive rats into five groups, including control. Give the rats in Group 1, methionine or met group, a diet of 3%methionine for 28 days to induce H-type hypertension. For Group 2 to 4 rats, administer a combination of Huotan Jiedu Tongluo Decoction and enalapril maleate in addition to the methionine diet for 28 days.Next, create a restraining device by placing a cylindrical restraint mesh into a thermal tube and putting the thermal tube into a canvas cover. After 28 days of treatment, place a rat in the mesh. Next place the signal cable of a noninvasive sphygmomanometer under the cover.Position the rat's tail in the gap in the cover and secure it on the stabilizing form pad. Transfer the rat to a quiet measurement site 20 to 30 minutes before blood pressure measurement. Now connect the pressurized sensor's air hose connection, signal connection, and holding tube.Place the pressurization sensor at the tip of the rat's tail. A pulse wave should appear after the rat tail is inserted into the sensor. Press Start/Stop to initiate or stop the measurement.When the blood pressure test is completed, the result menu will pop up automatically. Check the average value of the measurement, standard deviation, standard, and coefficient of variation in the results menu. After euthanizing the rat, dissect it with a pair of surgical scissors and tooth forceps from the perineum to the neck.Turn over the thoracic and abdominal contents. Then navigate to the abdominal aortic bifurcation between the two kidneys. Collect the aorta up to a section of the aortic arch.The systolic and diastolic blood pressures were significantly greater in the Met Group relative to the control group from one to four weeks. The blood pressures decreased after the administration of the Huotan Jiedu Tongluo Decoction. The combined effect of the decoction and/or enalapril maleate had a stronger anti-hypertensive effect than the decoction alone.

Metiyonin ile İndüklenen H-Tipi Hipertansif Sıçanların Huotan Jiedu Tongluo Deoction (HTJDTLD) ile Tedavisi

To begin, centrifuge the coagulated blood-containing tubes at 626 to 1, 409 G for 20 minutes at two to eight degrees Celsius. Then carefully collect the supernatant in a new tube. Store the supernatant at minus 80 degrees Celsius.Next, set up blank standard and sample wells. Pipette 50 microliters of the diluted standards into the ELISA plate. Then transfer 40 microliters of the diluted samples into the sample wells.Add 10 microliters of serum into the sample wells. Now, seal the plate with a sealing film. Place the sealed plate in an incubator at 37 degrees Celsius for 30 minutes.Dilute the 30 times concentrated wash solution with distilled water to obtain a single strength wash solution. Next, remove the sealing film from the plate and discard the liquid. Now, fill each well with 200 microliters of washing solution five times.After discarding the supernatant for the last time, tap the plate dry. Pipette 50 microliters of the enzyme reagent into all wells other than the blank ones. Then seal the plate with a sealing film before placing it in an incubator at 37 degrees Celsius for 30 minutes.After washing the plate five more times, add 50 microliters of each of the color developers, A and B.Shake the plate gently to mix well. Then incubate the plate at 37 degrees Celsius at low light for 10 minutes. Now, pipette 50 microliters of the stop solution into each well to terminate the reaction.Measure the absorbance of each well sequentially at 450 nanometers. The plasma homocysteine concentration in the methionine group was twofold greater than in the control group. The plasma homocysteine concentrations were relatively lower in the treatment groups.