Method for diagnosing hepatitis transit into chronic stage

FIELD: medicine.

SUBSTANCE: method involves determining collagenase, collagenase inhibitor, protease inhibitor activity, free oxyproline and protein-bound oxyproline content. Collagenase activity increasing to 2.2±0.2 mcmole/l*h, protease inhibitor activity falling to 19.2±0.5 IU/ml and lower, collagenase inhibitor content falling to 2.1±0.2 IU/ml and lower, free oxyproline content growing to 1.2±0.02 IU/ml and protein-bound oxyproline content growing to 11.7±0.7 IU/ml, I stage of chronic transition is to be diagnosed. Collagenase activity increasing to 3.52±10.06 mcmole/l*h, collagenase inhibitor activity and protease inhibitor activity and free oxyproline content remaining the same, II stage of chronic transition is to be diagnosed. Collagenase activity increasing to 3.5±0.06 mcmole/l*h, protease inhibitor activity falling to 10.6±0.1 IU/ml and free oxyproline content growing to 1.524±0.06 IU/ml, III stage of chronic transition is to be diagnosed. Collagenase activity increasing to 4.06±0.02 mcmole/l*h, protease inhibitor activity falling to 9.0±0.28 IU/ml and lower, collagenase inhibitor content falling to 1.4±0.1 IU/ml, and protein-bound oxyproline content growing to 12.75±0.1 IU/ml, IV stage of chronic transition is to be diagnosed.

EFFECT: high accuracy of prognosis.

1 tbl

 

The invention relates to the field of medicine, Hepatology and can be used to diagnose the stage of chronic hepatitis.

A known method for the diagnosis of the stage of chronic hepatitis on histological grounds (V.Desmet et al, 1994), [11] which determine the nature of severity of fibrotic changes, and thereby assess the stage of chronicity of the process.

The known method is invasive with biopsy of the liver taking of biological material and then hold him histological examination with the calculation of the index of activity (IGA) and histological fibrosis index (HYPHAE).)

However, the stage of chronic hepatitis reflects a temporary its course and is characterized by the degree of liver fibrosis until the development of cirrhosis. Cirrhosis is considered as irreversible stage of chronic hepatitis.

A new technical result is to increase the accuracy and informativeness by reducing the number of complications.

The task to solve a new way to diagnose the stage of chronic hepatitis in the liver, consisting in the patient study, and investigate the serum with the determination of the activity of collagenase, collagenase inhibitor, proteasome inhibitor, the amount of free hydroxyproline and protein-bound hydroxyproline and with increasing activity call genasi to 2.2± 0.2 µmol/l·h, reducing the activity proteasome inhibitor to 19.2±0,5 S/ml and below, the activity of the collagenase inhibitor to 2.1±0,2 S/ml and below, increasing the content of free hydroxyproline to 1.2±0,02 S/ml, and protein-bound hydroxyproline to 11.7±0,7 S/ml diagnosed stage I chronic, while enhancing the activity of collagenase to 3.52±0.06 µmol/l·h, the increase in the content of free hydroxyproline to 1.45±0,02, at the same level of activity inhibitor of collagenase activity and protease inhibitor content of protein-bound hydroxyproline diagnosed with stage II chronic, increasing the activity of collagenase to 3.5±0.06 µmol/l·h, reducing the activity proteasome inhibitor to 10.6±0,1 IE/ml, the increase in the content of free hydroxyproline through 1.52±0,01 S/ml, diagnosed with stage III chronic, increasing the activity of collagenase to 4,06±0.02 µmol/l·h, reducing the activity proteasome inhibitor to 9.0±0,28 S/ml and below, and inhibitor of collagenase to 1.4±0,1 IE/ml, the increase in the content of protein-bound hydroxyproline to 12.75±0,1 S/ml diagnosed with stage IV chronic.

The method is realized in the following way.

In the serum of the patient determine the activity of collagenase, collagenase inhibitor, proteasome inhibitor, the amount of free hydroxyproline and Belk is posvyathennogo hydroxyproline and by increasing the activity of collagenase to 2.2± 0.2 µmol/l·h, reducing the activity proteasome inhibitor to 19.2±0,5 S/ml and below, an inhibitor of collagenase to 2.1±0,2 S/ml and below, increasing the content of free hydroxyproline to 1.2±0,02 S/ml, and protein-bound hydroxyproline to 11.7±0,7 S/ml diagnosed stage I chronic, while enhancing the activity of collagenase to 3.52±0.06 µmol/l·h, the increase in the content of free hydroxyproline to 1,45±0,02 was diagnosed with stage II chronic, increasing the activity of collagenase to 3.5±0.06 µmol/l·h, reducing the activity of the inhibitor proteasome to 10.6±0,1 IE/ml, the increase in the content of hydroxyproline free to 1,52±0,01 S/ml diagnosed with stage III chronic, increasing the activity of collagenase to 4,06±0.02 µmol/l·h, reducing the activity of the inhibitor proteasome to 9.0±0,28 S/ml and below, and the activity of the collagenase inhibitor to 1.4±0,1 IE/ml, the increase in the content of protein-bound hydroxyproline to 12.75±0,1 S/ml diagnosed with stage IV chronic.

The proposed criteria are selected on the basis of analysis of data from clinical trials, 60 patients with chronic hepatitis of various stages of chronicity of the process in the liver, which were conducted histological examination of liver biopsy specimens and determination of biochemical parameters in the serum according to the proposed method

So, in those patients where fibrosis was absent completely the activity of collagenase in serum was not higher than the 12.7±0.04 µmol/l·h, activity proteasome inhibitor was not below 26,4±0,5 S/ml, collagenase inhibitor was not below 3,86±0,01 S/ml, the content of free hydroxyproline was not above 0,6±0,02 S/ml, and protein-bound no higher than 6,7±0,01 S/ml.

In patients with stage I fibrosis increased activity of collagenase - 2,2±0.2 µmol/l·h, decreased activity proteasome inhibitor below 19,2±0,5 S/ml, collagenase inhibitor decreased below 2.1±0,2 S/ml, increased the content of hydroxyproline free to 1.2±0,02 S/ml and protein-bound hydroxyproline to 11.7±0,7 S/ml.

In a phase II fibrosis activity of collagenase was increased to 3.52±0.06 µmol/l·h increased the content of hydroxyproline free to 1.45±0,02. The remaining indicators were not changed.

In the third stage of fibrosis activity of collagenase was increased to 3.5±0.06 µmol/l·h, decreased activity inhibitor proteasome to 10.6±0,1 IE/ml, increased the content of hydroxyproline free to 1,52±0,01 S/ml. Other indicators did not change.

At stage IV chronic (cirrhosis) activity of collagenase was growing up 4,06±0.02 µmol/l·h, decreased activity inhibitor proteasome to 9.0±0,28 S/ml and below, the activity inhibitor is and collagenase was decreased to 1.4± 0,1 S/ml, increased the content of protein-bound hydroxyproline to 12.75±0,1 S/ml. Other indicators remained the same.

Apparently the identified correlation is attributable to the following: biodegradation of collagen carried out by a specific protease-collagenase [4, 6, 7], can be regarded as the activation system collagenolysis in response to increased synthesis of collagen [3]. At the same time, the reduced activity of inhibitory link - reduced activity proteasome inhibitor and an inhibitor of collagenase [1]. Degradation of collagen leads to increased levels of hydroxyproline and its fractions - free hydroxyproline and protein-bound hydroxyproline [10].

According to biopsy of the liver, proving the existence of a certain stage of fibrosis sootvetstvenno stage of chronicity of the process and parallel biochemical studies of blood on the activity of collagenase [8], inhibitors of collagenase and proteases inhibitor [5], and determining the content of hydroxyproline - its fractions free hydroxyproline and protein-bound hydroxyproline [2] can be judged on existing changes of biochemical parameters of blood, associated with increasing stage of fibrosis.

When I stage of fibrosis was significantly increased (almost 2 times - 2,2±0,04) collagenase activity, decreased activity of the collagenase inhibitor (p<0,05) (almost 2 the Aza - 2,0±0.2) and 1.5 times decreased activity proteasome inhibitor (19,2±0.5), and increased in 2 times the content of hydroxyproline free and protein-bound (p<0,05), respectively 1,2±0.02 and 11,7±0,7 S/ml.

In stage II fibrosis activity of collagenase increases (3 times - 3,2±0,06), increases the content of free hydroxyproline (p<0,05) more than 2 times - 1,45±0,02 S/ml.

In the third stage of fibrosis increases the activity of collagenase (3,5±0,06), reduced activity proteasome inhibitor 10,6±0,19 (almost in 2 times in comparison with stage I), increases the content of free hydroxyproline in 2 times - 1,52±0,01 p<0,05) S/ml.

Example 1. Patient 39 years, the diagnosis of chronic hepatitis of viral etiology, type, stage I fibrosis, moderate stage of activity, HBcor AgIgG+, HBsAG+, increased Tran-samines - 2 times, the liver was enlarged to 1.5 cm below costal arch, painless, soft, positive PCR for HBV DNA. For biochemical analysis of blood disorders belkovosinteticescuu liver function were observed, prothrombin index was 90. Study according to the proposed method. For biochemical analysis of blood activity of collagenase was 3.2 µmol/l·h, proteasome inhibitor - 19,0 IE/ml, the activity of the collagenase inhibitor was reduced to 2.08, increases the content of hydroxyproline free to 1.25 S/ml, belkovo is related hydroxyproline to 10.0 S/ml The obtained data allowed the diagnosis of chronic hepatitis stage I chronic (stage I fibrosis).

Subsequent histological examination of the liver diagnosis confirmed.

Example 2. Patient 32 years old, diagnosed with chronic hepatitis of viral etiology, type in the replication phase (positive PCR for HCV RNA), stage II fibrosis. For biochemical analysis of blood transaminases were normal, was normal albumin levels and prothrombin index, the activity of collagenase increased to 3.58 μmol/l·h, the content of free hydroxyproline was increased to 1.43 S/ml. was diagnosed as chronic hepatitis, II stage of chronicity.

Histological examination of the liver, the diagnosis was confirmed.

Example 3. Patient, 36 years of age, the diagnosis of chronic hepatitis of viral etiology, type in the replication phase of viral infection, minimal activity, III stage of fibrosis. The patient had no complaints, liver even royalty 1 cm below the costal arch, painless, plotnovato. It was noted peripheral hepatic signs in the form of a single telangiectasias. For biochemical analysis of blood transaminases were increased slightly, did not exceed more than 1.5 times, the content of albumin was within bromeliad - 38 g/l, the value of the prothrombin index was equal to 85. According to additionally the studies according to the proposed method - increased activity of collagenase to 3.56 mmol/l·h, proteasome inhibitor decreased to 11.6 S/ml, the content of free hydroxyproline was increased to a rate of 1.51 S/ml: diagnosed chronic hepatitis fibrosis stage III, which were affirmed biopsy of the liver

Example 4. Patient C., 49 years old, diagnosed with liver cirrhosis of viral etiology, the minimum stage of activity. In biochemical studies transaminases were within normal limits, the bilirubin concentration was 20 mg/%, of which direct and 12 indirect - 8, albumin was 35 g/l, prothrombin index - 82, further study was conducted according to the proposed method, the activity of the collagenase increased to 4.04 mmol/l·h, activity proteasome inhibitor decreased to 8,72 IE/ml, collagenase inhibitor decreased to 1.5 IE/ml, the content of protein-bound hydroxyproline was increased to 12,74 IE/ml, the diagnosis of stage IV fibrosis, cirrhosis was confirmed data biopsy of the liver.

Thus, using the proposed method with high accuracy and information content to diagnose the stage of the chronicity of the liver in patients with hepatitis, avoiding the use of invasive way associated with biopsy of the liver that in some stages of the disease are contraindicated, as it may cause bleeding.

The literature dealing with the RA

1. Zorin N.A. study of the reactions between proteinase blood plasma and collagenase / Nasirin, Sgjoen, Irgizla // Questions of medical chemistry. - 1993. - V.41, №6. - P.53.

2. Kuznetsova ETC., Proshina L.Y., Prevalence LI modification of the definition of the content of hydroxyproline in serum // Laboratory work. - 1982. No. 8. - P.8-10.

3. Logvinov A.S., Block J.E. Chronic hepatitis and cirrhosis of the liver. - M.: Medicine, 1987.

4. Mackevicius Z.K. Mechanisms and the role of biodegradation of collagen in pathology / Schmatics // Archives of pathology. - 1987. - V.49, no. 6. - P.3-9.

5. Artikova V.F., Pashina FORCE Uniform method for determining the activity α1antitrypsin and α2macroglobulin in the serum (plasma) human blood // the Questions of medical chemistry. - 1979. - V.25, No. 4. - S-499.

6. Serov V.V. connective tissue / CEO, Abecher. - M.: Nauka, 1981. - 456 S.

7. Slutsky LI Biochemistry of normal and pathological changes of the connective tissue / Lieselotte. - M.: Nauka, 1969. - 345 S.

8. Sharayev P.N., Peskov V.N., Zvorykina N.G. Definition collagenolytic activity of blood plasma // Laboratory work. - 1987. - No. 1 - P.60-62.

9. Sharayev I.N. Method for the determination of free and bound hydroxyproline in serum // Laboratory work. - 1981. No. 5. - S-285.

10. Urbanec E.A. the Effect of collagenase on neuromuscular transmission frogs / EA Urkan the C // Physiological journal of the USSR Sechenov. - 1987. - T, 34. - S-491.

11. Desmet V., Gerber M., Hoffnagle H. et al. Classification of chronic hepatitis, diagnosis, determine the severity and gravity of the current // Russian journal of gastroenterology, Hepatology and Coloproctology. - 1995. No. 2. - Ñ.38-45.

Table 1

The stage of chronic hepatitis on histological characteristics - degree of fibrosis (histologic index (GI fibrosis))
GIThe degree of fibrosisMorphological features
0nono
1weakfibrosis and enlargement of portal tracts
2moderateperiportal fibrosis
3heavyorthocentrinae septa
4cirrhosiscirrhosis

The way to diagnose the stage of chronic hepatitis in the liver, consisting in the study of a patient, wherein investigate the serum with the determination of the activity of collagenase, collagenase inhibitor, proteasome inhibitor, the content of free hydroxyproline and protein-bound hydroxyproline and by increasing the activity of collagenase to (2,2±0,2) µmol/l·h, SN the terms activity proteasome inhibitor to (19,2± 0,5) S/ml and below, the activity of the collagenase inhibitor to (2,1±0,2) S/ml and below, increasing the content of free hydroxyproline to (1,2±0,02) IE/ml, and protein-bound hydroxyproline to (11,7±0,7) S/ml diagnosed stage I chronic, while enhancing the activity of collagenase to (3,52±0,06) µmol/l·h, the increase in the content of free hydroxyproline to (1,45±0,02) S/ml with the same level of activity inhibitor of collagenase activity and protease inhibitor content of protein-bound hydroxyproline diagnosed with stage II chronic, increasing the activity of collagenase to (3,5±0,06) µmol/l·h, reducing the activity proteasome inhibitor to (10,6±0,1) S/ml, the increase in the content of free hydroxyproline to (1,52±0,01) S/ml diagnosed with stage III chronic, increasing the activity of collagenase to (4,06±0,02) µmol/l·h, reducing the activity proteasome inhibitor to (9,0±0,28) IE/ml and below, and inhibitor of collagenase to (1,4±0,1) S/ml, the increase in the content of protein-bound hydroxyproline up (was 12.75±0,1) S/ml diagnosed with stage IV chronic.



 

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