How to count the analysis of peritoneal fluid. Modern problems of science and education. Diseases in which there is free fluid in the space of Douglas
Peritoneal dialysis is a method of artificial purification of blood from toxins, based on the filtration properties of the patient's peritoneum.
The peritoneum is a thin membrane that completely or partially covers the internal organs of the abdominal cavity. In physical terms, the peritoneum is a membrane with selective permeability for various substances. The peritoneum has three types of pores: small, water-permeable, medium, for the passage of water-soluble compounds and substances with a small molecular weight, and large - for substances with a large molecular weight. Due to its large penetrating ability, the peritoneum is able to pass various types of toxins. This distinguishes the method of peritoneal dialysis from hemodialysis, in which only substances with a small and partially medium molecular weight pass through the membrane.
In peritoneal dialysis, the dialysis solution (dialysate) is located in the abdominal cavity and constantly filters toxins from the vessels in the peritoneal wall. Within a few hours, the dialysate becomes contaminated with toxins, the filtration process stops, which requires the replacement of the solution.
The speed and volume of filtration is constant, the cleaning process is slow and long, which allows the use of peritoneal dialysis in patients with low or unstable blood pressure and in children. In addition to filtration during peritoneal dialysis, excess fluid enters the solution. This process is called ultrafiltration. The dialysate contains an osmotic active substance, such as a concentrated glucose solution, which attracts liquid along the concentration gradient. As a result, excess fluid from the bloodstream through the vessels of the peritoneum enters the dialysis solution. In addition to glucose, amino acids, dextrose, glycerol, starch are present as an osmotic agent in some dialysis solutions. In addition, the dialysate contains a complex of chemicals, selected depending on the needs of the patient.
Indications for peritoneal dialysis
Peritoneal dialysis is preferable to hemodialysis in the following cases:
For patients in whom it is not possible to create adequate vascular access (persons with low blood pressure, severe diabetic angiopathy, young children).
For patients with severe diseases of the cardiovascular system, in whom hemodialysis sessions can lead to the development of complications.
For patients with bleeding disorders, in whom the use of drugs that prevent thrombosis is contraindicated.
For patients with intolerance to synthetic hemodialysis filter membranes.
For patients who do not want to depend on a hemodialysis machine.
Contraindications for peritoneal dialysis
Peritoneal dialysis is contraindicated in:
The presence of adhesions in the abdominal cavity, as well as an increase in internal organs, which limits the surface of the peritoneum.
With established low filtration characteristics of the peritoneum.
The presence of drainage in the abdominal cavity in adjacent organs (colostomy, cystostomy).
Purulent diseases of the skin in the abdominal wall.
Mental illness, when the patient is not able to properly conduct a session of peritoneal dialysis.
Obesity, when the effectiveness of blood purification in peritoneal dialysis is questioned.
peritoneal dialysis procedure
The kit for peritoneal dialysis includes containers (empty and with solution) and conductive lines.
Cyclers are also used during the procedure. The cycler is a device that provides programmable cycles for filling and draining the solution, as well as being able to heat the solution to the desired temperature and weigh the drained dialysate to estimate the volume of fluid removed.
Peritoneal catheters are used to access the abdominal cavity.
Catheters should provide good drainage of the abdominal cavity, be tightly fixed, and be protected from infection. Adequate irrigation of the abdominal cavity is carried out due to the high speed of filling and draining the solution. The catheter is tightly fixed in the subcutaneous fat due to the germination of the Dacron cuff with connective tissue. It also creates a barrier to infection. Catheters are made of silicone or polyurethane. The catheter is placed surgically into the pelvic cavity. The outer part of the catheter is brought out under the skin on the anterior or lateral surface of the abdominal cavity.
After placing the catheter for adequate fixation, 2-3 weeks should pass, after which they begin to conduct dialysis sessions.
For peritoneal dialysis, it is necessary to attach a container filled with dialysis solution to the catheter.
This process takes place subject to hygienic and antiseptic rules, including the treatment of hands, work surfaces, the skin around the catheter, as well as the junctions of the lines and the catheter (adapter), putting on a mask on the face. The front surface of the abdomen is freed from clothing, a clean cotton towel is tied to the belt. From the sterile bag, a drain empty bag and a container with fresh dialysis solution are taken out. In this case, the container with fresh solution is hung on a tripod at a height of 1.5 m, and the drain bag is placed on the floor. The mains after treatment with an antiseptic solution are interconnected.
First, the solution is drained into an empty bag. Then this part of the highway is pinched, the clamp opens on the bringing branch of the highway. New dialysis fluid is poured into the abdominal cavity. After that, the clamps on the lines are clamped, the empty container and the bag with the drained solution are removed. The outer port of the catheter is closed with a protective cap, fixed to the skin and hidden under clothing. Every month, patients take blood and fluid from the abdominal cavity for examination. Based on the results, a conclusion is made about the degree of blood purification, as well as the presence or absence of anemia, disorders of phosphorus-calcium metabolism, and, based on these indicators, the treatment is corrected. On average, exchange sessions are carried out 3 times a day, the volume of dialysis solution is 2-2.5 liters.
In case of poor tolerance, non-compliance with the regimen, insufficient blood purification, as well as in the event of severe or recurring complications, it is recommended to transfer the patient to hemodialysis.
Complications of peritoneal dialysis
The most dangerous complication of peritoneal dialysis is peritonitis (inflammation of the peritoneum). The most common reason for the development of inflammation is the patient's failure to follow the rules of antiseptics during exchange sessions. Peritonitis is diagnosed when two of three signs are present:
External manifestations of inflammation of the peritoneum: pain in the abdomen, fever, chills, general weakness, nausea, vomiting, impaired stool.
Turbid peritoneal fluid.
Detection of bacteria in peritoneal fluid.
Treatment: broad-spectrum antibiotics until the test results, then an antibacterial drug, taking into account the sensitivity of the identified microorganisms to it. In addition to specific therapy, a temporary cessation of peritoneal dialysis sessions, washing of the abdominal cavity with a standard dialysis solution or Ringer's lactate solution is recommended. Heparin is added to the solutions during washing, which prevents the adhesive process in the abdominal cavity. In severe cases, removal of the peritoneal catheter may be necessary.
Non-infectious complications include the following:
Violation of the abdominal catheter with difficulty filling / draining the solution. This complication may be due to a change in the location of the catheter, closure of the catheter by a loop of intestine, for example, with constipation, bending of the catheter, or closure of the lumen of the catheter with blood clots or fibrin, which is often found in peritonitis. When closing the lumen of the catheter with clots, you can try to flush it with a sterile isotonic solution. If unsuccessful, catheter replacement is indicated. Complications associated with changing the position of the catheter require surgical intervention.
When the gulf is found and the dialysis solution is in the abdominal cavity, intra-abdominal pressure increases, which contributes to the formation of hernias. The most common hernias of the white line, less often umbilical and inguinal hernias. Depending on the size and reducibility of the hernial protrusion, the question of further treatment is decided: surgery or expectant management.
The outflow of peritoneal solution outward or into the subcutaneous fatty tissue occurs, as a rule, immediately after the placement of an intra-abdominal catheter, or with poor fixation of the catheter in elderly and debilitated patients. This complication is diagnosed when the bandage gets wet in the area of the catheter, or when edema of the subcutaneous fat of the abdominal wall and genitals forms. Treatment consists in stopping peritoneal dialysis for 1-2 weeks for optimal fixation of the catheter, with the patient undergoing hemodialysis sessions. Under adverse conditions, catheter replacement is indicated.
Right-sided pleurisy occurs in debilitated patients, as well as in some patients at the beginning of treatment. This complication is associated with the penetration of dialysis fluid through the diaphragm into the pleural cavity. Treatment - reducing the volume of the poured solution. To prevent this condition, it is recommended to conduct exchange sessions in a vertical state. With an increase in respiratory failure, the transfer of the patient to program hemodialysis is indicated.
Abdominal pain that is not associated with inflammation of the peritoneum often occurs at the beginning of treatment and disappears after a couple of months. The pain is usually associated with irritation of the peritoneum by a chemically active dialysis solution, or due to overdistension of the abdominal cavity with a large amount of solution. In the first case, the treatment consists in the selection of the dialysate that is optimal in chemical composition, in the second case, the filling of smaller volumes of solutions with an increase in the frequency of exchanges.
Many experts consider peritoneal dialysis as the first stage of replacement therapy for patients in the terminal stage of renal failure. In some patients, for a number of reasons, peritoneal dialysis is the only possible treatment.
Compared with hemodialysis, peritoneal dialysis allows patients to lead an active lifestyle and work. But, unfortunately, the duration of treatment with peritoneal dialysis directly depends on the filtering properties of the peritoneum, which, over time, gradually, and with frequent peritonitis rather quickly, decrease. In this case, there is a need for alternative methods: hemodialysis or kidney transplantation.
Therapist, nephrologist Sirotkina E.V.
Normal serous fluid of the abdominal cavity is clear and light yellow, the volume is less than 50 ml. If fluid in the abdominal cavity accumulates in a significant amount, it is called ascitic. Ascites - abdominal dropsy, dropsy of the abdomen, a significant accumulation of free fluid (usually transudate) in the abdominal cavity. Ascites may occur suddenly (for example, with portal vein thrombosis) or develop gradually over several months, accompanied by flatulence, which may initially dominate the clinical picture. Sometimes in the peritoneal cavity accumulates from 8 to 30 liters of ascitic fluid. During a physical examination of a patient, ascites can be recognized if there is at least 1 liter of fluid in the peritoneal cavity.
OBTAINING AND PROCESSING OF MATERIAL FOR LABORATORY STUDIES
The serous fluid of the abdominal cavity is obtained by puncture of the recto-uterine recess (Douglas space, pocket) (culdocentesis), by percutaneous puncture (paracentesis) or by laparoscopy.
At the same time, 5 ml of venous blood should be taken to determine the serum-ascitic fluid gradient for albumin and other biochemical parameters.
Cytological examination of ascitic fluid is desirable to be carried out immediately after delivery of the sample to the laboratory. If emergency analysis is not possible, the sample should be stored in a refrigerator for no more than 12 hours using heparin or sodium citrate as an anticoagulant.
CLINICAL AND DIAGNOSTIC SIGNIFICANCE OF CHANGES IN THE INDICATORS OF SEROUS FLUID IN THE ABDOMINAL CAVITY
The main tasks of the laboratory analysis of ascitic fluid are:
establishment of a benign or malignant nature of the effusion;
differentiation of non-infection/infection of the fluid.
Ascites in portal hypertension, hepatocellular carcinoma and liver metastases without spreading through the peritoneum is a transudate.
Ascites in liver diseases, in cases of pancreatitis, peritoneal tuberculosis and in malignant tumors with metastases in the peritoneum is usually the result of exudate.
Appropriate laboratory criteria have been developed for the differential diagnosis of portal and malignant (peritoneal metastases) types of ascites. The main laboratory indicators of the separation of ascites into exudate and transudate in the absence of malignant cells are the content of albumin, the level of cholesterol and fibronectin. Transudate is characterized by a high albumin gradient between serum and ascitic fluid (> 11 g/l), for exudate, on the contrary, ascitic fluid contains a large amount of albumin and the gradient for albumin between ascitic fluid and serum is insignificant (
In the case of chylous ascitic fluid, the determination of triglycerides in it and the setting of lipoprotein electrophoresis are indicated. The level of cancer-embryonic antigen of more than 2.5 μg/l of ascitic fluid has a high clinical specificity and a prognostic value close to 100% for a tumor with peritoneal metastases.
In the study of albumin, serum and ascitic fluid samples should be taken at the same time. The concentration of albumin in ascitic fluid should be determined by immunonephelometric or immunoturbidimetric method. Photometric determination of albumin with bromcresol green gives overestimated results at albumin concentrations of more than 7 g/l, so this method is not suitable for determining the albumin gradient. A total protein level of more than 30 g/l for the diagnosis of transudate has a diagnostic specificity of 86% and a diagnostic specificity of 83%.
Cell counting and differentiation to determine neutrophilic granulocytes is carried out in EDTA-ascitic fluid. If blood is present in the sample, then it is necessary to determine:
the ratio of erythrocytes to leukocytes to assess the possibility of bleeding from the gastrointestinal tract or the admixture of "travel" blood;
the content of leukocytes (their high relative number indicates an inflammatory process);
the content of neutrophilic granulocytes (ascites with a predominance and an absolute content of neutrophils more than 250/µl is classified as infectious).
If a cytological examination cannot be performed within 12 hours, the sample can be stored in the refrigerator for up to 2 days, however, the material should be fixed by adding 50% alcohol in a ratio of 1:1.
The determination of pathogenic microflora in ascitic fluid is carried out in the same way as in the blood - by cultivation under aerobic and anaerobic conditions.
GENERAL PROPERTIES (MACROSCOPIC VIEW OF LIQUID)
Ascitic (peritoneal) fluid by its nature is more often serous, less often hemorrhagic, chylous, mucous. Light, transparent or yellowish ascitic fluid is more often a transudate. Cloudy serous fluid is characteristic of peritonitis, which occurs as a complication of appendicitis, pancreatitis, intestinal obstruction, and primary bacterial infection. Greenish color, bile staining occurs with perforation of the gallbladder, duodenal bulb, small intestine, as well as with cholecystitis, acute pancreatitis. Ascitic fluid acquires a greenish color when the concentration of bilirubin in it is more than 100 μmol / l. If the concentration of bilirubin in ascitic fluid is higher than in serum, this is evidence of perforation of the bile duct or bladder. The milky appearance of the serous fluid (chylous effusion) appears when there are a large number of chylomicrons in the ascitic fluid. This happens most often with damage or obstruction of the thoracic lymphatic duct caused by tuberculosis, cirrhosis of the liver, and can occur with leukemia (lymphoma). Pseudochile infusion is possible when large volumes of blood substitutes are administered to patients.
Hemorrhagic serous fluid of the abdominal cavity may appear with abdominal trauma with rupture of internal organs, in particular, with rupture of the fallopian tube during ectopic pregnancy, as well as with dissemination of a malignant tumor along the serous membrane. The liquid has a scarlet color with the admixture of "travel" blood during paracentesis. Ascitic fluid acquires a brown color due to bleeding into the abdominal cavity with tuberculous peritonitis, metastases to the peritoneum and after traumatic injuries of the abdominal organs. In the integral assessment of the condition of patients with peritonitis, the Mannheim peritoneal index is used, in the calculation of which one of the most significant indicators is the nature of the exudate.
BIOCHEMICAL INDICATORS
The concentration of total protein in ascitic fluid is mainly influenced by the following factors:
the concentration of total protein in blood serum, with which the level of protein in ascitic fluid has a direct relationship;
the level of portal hypertension, with which there is an inverse relationship. In addition, the concentration of total protein in ascitic fluid is affected by
taking diuretics.
Indications for the determination of total protein in ascitic fluid are:
prophylactic antibiotics to prevent bacterial peritonitis;
differential diagnosis of primary and secondary (introduced) bacterial peritonitis;
ascites in heart failure.
total protein
With a threshold value of the total protein content in the ascitic fluid of 25 g/l, the classical ideas about the separation of transudate and exudate based on protein exudation are confirmed only by 56%, i.e., they practically do not have laboratory confirmation. This is due to the fact that the content of total protein is low in the infectious etiology of ascites, although the exudative nature of ascites is undeniable. On the other hand, the content of total protein in the ascitic fluid is high in patients with heart failure, in which the ascitic fluid is considered as a transudate.
A total protein concentration greater than 30 g/l indicates an exudate with a clinical sensitivity of 93% and a specificity of 85%. A protein ratio value greater than °-5 indicates exudate with a clinical sensitivity of 93% and a specificity of 85%. In ascites caused by malignant tumors, with the exception of hepatocellular carcinoma, the concentration of total protein is 49-65 g/l, while in liver cirrhosis and hepatocellular carcinoma it is in the range of 17-21 g/l. Patients with liver cirrhosis and protein levels in ascitic fluid less than 15 g/l have a poor prognosis. A low level of protein in the ascitic fluid is characteristic of the infectious nature of ascites, while at the same time, with a secondary bacterial infection and with tuberculous peritonitis, the protein level of more than 30 g / l is constantly determined in the ascitic fluid.
Albumen
The albumin gradient between serum and ascitic fluid is determined by the level of portal hypertension. Patients with an albumin gradient greater than 11 g/l have portal hypertension, while patients with an albumin gradient less than 11 g/l do not. In cirrhosis of the liver with portal hypertension, an albumin gradient of more than 11 g/l has a clinical specificity of 97%. Ascites in mixed types - due to cirrhosis of the liver and peritoneal metastases or cirrhosis of the liver and tuberculous peritonitis - are also accompanied by an albumin gradient of more than 11 g / l.
To distinguish transudate from exudate, the albumin gradient is more important than total protein. It should be borne in mind that when using diuretics or paracentesis, the indicators of albumin and total protein in ascitic fluid change.
In patients with widespread peritonitis, the degree of decrease in serum albumin concentration is a highly informative prognostic indicator that allows assessing the severity of the disease and the risk of an adverse outcome. A decrease in the level of albumin in the blood occurs both due to an increase in protein catabolism, which is characteristic of the acute phase of any inflammatory process, and due to exudation into the abdominal cavity. Peritoneal exudate contains a significant amount of protein. It is believed that up to 50% of the total extracellular fluid of the body can move into the abdominal cavity. With the development of edema, at first the process of fluid absorption by the peritoneum even accelerates, but then, due to a violation of microcirculation, it slows down sharply - exudate accumulates. Some amount of protein, including albumin, released from the bloodstream, is lost to the body; High protein effusion leads to hypoalbuminemia. In addition, there is always a significant swelling of the pre- and retroperitoneal tissue and other tissues due to the action of biologically active substances: enzymes, kinins, histamine, due to which albumin is deposited in the tissues. Along with a decrease in synthesis (albumin is a negative reactant of the acute phase of inflammation) and an increase in decay (the predominance of catabolism processes in the body), the release of albumin into the abdominal cavity and its deposition in edematous tissues are the main cause of a prognostically unfavorable phenomenon - a decrease in the concentration of albumin in blood serum.
The penetration of albumin from the blood into the effusion is characterized by the value of the total concentration of albumin in the effusion. In most cases, the total concentration of albumin in the effusion is in close correlation with the value of the total concentration of albumin in the effusion (the concentration of albumin in the blood serum). In cases where the total concentration of albumin in the effusion is >34 g/l and the total concentration of albumin in the effusion is significantly lower than the total concentration of albumin in the effusion, patients have a mild postoperative period; the outcome of the disease is favorable; discharge through the drains is insignificant.
In cases of a significant difference in the state of albumin in the effusion and serum, the value of the total concentration of albumin in the effusion is not related to the concentration of albumin in the blood; obviously, there is a proteolytic effect on albumin in the exudate, leading to its change and destruction. In other cases, the absolute value of the total concentration of albumin in the effusion depends on the value of the total concentration of albumin in the effusion, and not on the activity of the inflammatory process in the abdominal cavity. To characterize the severity of the inflammatory process in the abdominal cavity, the ratio of the total concentration of albumin in the effusion is suitable, which characterizes the degree of permeability of the vessel wall for albumin. The parameter of the total concentration of albumin in the effusion reflects the severity of the patient's condition, and the parameter of the total concentration of albumin in the effusion reflects the severity of the inflammatory process in the abdominal cavity.
Endotoxin
In patients with fecal peritonitis (perforation of the large intestine), the concentration of endotoxin in the peritoneal fluid can reach 1000 μg / l. In diffuse bacterial peritonitis, the concentration of endotoxin is usually relatively low. If in the postoperative period the microflora of the colon continues to enter the abdominal cavity (endotoxin in the peritoneal fluid increases), this is a very big threat of diffuse peritonitis and death of the patient.
Glucose
With non-bacterial peritonitis, the value of the ratio of glucose-ascitic fluid / glucose plasma is more than 1, with bacterial peritonitis, this value is less than 1. The concentration of glucose in the ascitic fluid is less than 2.8 mmol / l is characteristic of bacterial peritonitis. In most cases of tuberculous peritonitis, the concentration of glucose in the ascitic fluid is less than 1.7 mmol / l. It is also reduced in malignant tumors with metastases, which are the causes of ascites.
BACTERIOLOGICAL STUDY
When the peritoneal fluid is infected with a monoculture, cultural studies have a clinical sensitivity of more than 85-90%, while at the same time, such an indicator as the content of neutrophilic granulocytes / μl of more than 250 is only 50%. Bacteriological studies show that in 70% of cases there is a gram-negative microflora, usually E. coli, in 20% of cases - gram-positive.
The etiological significance of anaerobic microorganisms has been proven in all complicated intra-abdominal infections, where they are part of polymicrobial associations. Clostridium perfringens and Clostridium septicum are involved in the development of secondary peritonitis, intra-abdominal abscess and sepsis with perforation of the colon and terminal ileum. With a complicated tumor of the colon, 70-85% of patients had bacteremia caused by Clostridium septicum. In appendicitis, Bilophila wadsworthia is shed from the effusion. Mixed infections often involve Peptococcus spp., Peptostreptococcus spp. Occurring in 90-95% of cases, Actinomyces spp., Prevotella spp., Lactobacillus spp., Fusobacterium spp. most difficult in the diagnosis and treatment of actinomycotic liver abscesses. Bacteroides, primarily Bacteroides fragilis and Bacteroides thetaiotaomicron, play a leading role in intra-abdominal infections. The frequency of isolation of Bacteroides fragilis in secondary peritonitis reaches 22.8-44.5%. There is evidence of the etiological significance of anaerobes in primary peritonitis. Clostridium perfringens and Bacteroides fragilis are isolated from all patients with biliary tract infections with signs of biliary hypertension or endoprostheses providing patency of the common bile duct. In pancreatitis, bacteroids and clostridia are distinguished among pathogens in 5-14% of cases. In nosocomial intra-abdominal infection, the association of multiresistant enterobacteria, Pseudomonas aeruginosa, enterococci and anaerobes is significant. Wound infections in patients after abdominal interventions in 3.9% are caused by Bacteroides spp., in 1.1% - by Clostridium perfringens. The development of peritonitis and mortality in intra-abdominal infections are due to gram-negative aerobes, and the formation of intra-abdominal abscesses in survivors is due to anaerobic microflora.
The progressive microbial-inflammatory process in the abdominal cavity (against the background of an operating injury, a decrease in the body's resistance, etc.) proceeds very rapidly and within 4-5 days can reach a stage requiring surgical intervention. The dynamism of the process, the high probability of reinfection, the development of microflora resistance require both the empirical prescription of antibiotic therapy simultaneously with an indicative microbiological express analysis (microscopy, gas chromatography) and an assessment of the sensitivity of microorganisms in clinical material using disks with antibiotics, as well as constant monitoring of the bacteriological situation in hotbed of inflammation. The routine bacteriological method of cultivation allows obtaining the result 24-72 hours after the start of the study, revealing the nature of the pathogenic microflora, confirming the correct choice of antibiotic therapy or suggesting the need for its correction. Constant microbiological monitoring is necessary because the microbiological situation in the focus of infection can change significantly in the course of the pathological process and require a change in therapy.
BLOOD CELL STUDY
Erythrocytes in EDTA-ascitic fluid
Peritoneal lavage fluid contains less than 25,000 RBCs/µl. In ascitic fluid, erythrocytes accumulate after injuries or are a manifestation of tuberculous or malignant lesions of the peritoneum. A similar ratio of the number of erythrocytes and leukocytes in ascitic fluid and blood indicates an iatrogenic lesion ("travel" blood) or bleeding into the abdominal cavity.
Leukocytes
Determination of leukocytes in the peritoneal fluid can be carried out quantitatively by counting in the Goryaev chamber or semi-quantitatively - using diagnostic strips using a test zone for the determination of leukocytes. The sensitivity of the test zone for leukocytes is 3000 cells/µl. The diagnostic sensitivity of the detection of leukocytes by the method of "dry chemistry" in the peritoneal fluid is 88%, the specificity is 94%.
The number of leukocytes in ascitic fluid, pH and lactate concentration during infection of ascitic fluid differ significantly from the values of these indicators in sterile ascites, however, all these values change approximately the same way and it is not possible to distinguish between ascites in tumor and infectious peritonitis.
The relative content of neutrophilic granulocytes of more than 25% of the leukocyte population of ascitic fluid is considered pathological and characteristic of bacterial peritonitis. In patients with cirrhosis of the liver, the number of leukocytes in ascitic fluid is inversely proportional to the volume of fluid. The number of leukocytes in the ascitic fluid increases with a decrease in the volume of ascites as a result of treatment with diuretics, while the relative content of neutrophils in the leukocyte formula does not change.
If the leukocyte count is more than 500/µl, especially if the granulocytes are more than 250/µl, the likelihood of primary bacterial peritonitis is high. In this case, the diagnostic specificity of confirming this diagnosis is 93%, the sensitivity is 84%. In pancreatic cancer and hepatocellular cancer, moderate leukocytosis occurs - 300-1000 / μl. In alcoholic cirrhosis, the number of leukocytes in the ascitic fluid rises to 1100-21,000/µl.
Lymphocytosis in ascitic fluid is a sign of prolonged congestive exudation, chronic inflammation, tuberculosis, or a tumor process. In patients with cirrhosis of the liver and chylous ascitic fluid, the relative content of lymphocytes in it ranges from 12 to 96%, on average 70%.
Tumor markers
Carbohydrate antigen 19-9
In ascitic fluid at a threshold value (discriminant level) of 30 U/ml, the diagnostic sensitivity of a malignant tumor is 52%, the diagnostic specificity is 100%.
Cancer-embryonic antigen
The study of cancer-embryonic antigen in ascitic fluid makes it possible to differentiate benign and malignant diseases with a diagnostic specificity and sensitivity of 83%. With a threshold value of the cancer-embryonic antigen of 2.5 µg/ml, the diagnostic sensitivity is 45%, the diagnostic specificity is 100%. If the determination of the cancer-embryonic antigen in ascitic fluid is combined with the study of the presence of tumor cells, the diagnostic sensitivity in the case of a malignant tumor increases to 80%. It should be noted that the study of the cancer-embryonic antigen in ascitic fluid is of little significance for the diagnosis of diffuse malignant mesothelioma: the cancer-embryonic antigen is determined only in 1-9% of cases of this pathology. At the same time, the cancer-embryonic antigen is determined in 80% of cases of ascites associated with a tumor lesion in cancer of the stomach, breast, and lung.
CYTOLOGICAL STUDY
The highest analytical sensitivity is achieved with a cytological study of preparations stained according to Pappenheim and Leishman, and, if necessary, with an additional cytochemical study. The diagnostic sensitivity of detecting tumor cells under a light microscope is 40-70%. If it is possible to obtain cells from 200 ml or more of ascitic fluid, diagnostic sensitivity increases to 70-90% with almost 100% diagnostic specificity. Sometimes laboratory detection of tumor cells in ascitic fluid precedes the clinical manifestation of the tumor by 3 years or more.
In women, tumor cells in the ascitic fluid, in descending order, are more often found in tumors of the genital tract, especially the ovaries, then breast and gastrointestinal tract cancers. In men, dissemination in the peritoneum is most often determined with tumors of the gastrointestinal tract, leukemia. Approximately 80% of the tumors are adenocarcinomas. Of the pathological processes that are characteristic of lesions of the peritoneum and are practically not found in the pleura and pericardium, peritoneal pseudomyxoma should be noted.
Peritoneal involvement is seen with a mucus-producing tumor, most commonly a borderline ovarian mucinous tumor or appendix mucocele, as well as with well-differentiated ovarian or appendix mucinous adenocarcinoma. It is believed that the lesion can develop when a cystic tumor ruptures and generalized dissemination of tumor cells in the abdominal cavity. At the same time, a large amount of mucus accumulates in the abdominal cavity, which is difficult to aspirate due to its high density. Aspirate smears from pseudomyxoma of the peritoneum contain a large amount of positive mucus resistant to hyaluronidase. Tumor cells are usually few or not found in several preparations, so it is not uncommon to view several smears or repeat puncture to establish a diagnosis.
To establish the benign or malignant nature of the lesion, one should be very careful. With a benign lesion, the cells are small in size, with small dark nuclei, moderately abundant cytoplasm, and vacuoles are contained in the cytoplasm. The mucus often contains strands of fibrocytes. It cannot be concluded that a malignant process is based solely on the detection of a large amount of mucus, since it may be the result of a ruptured appendix mucocele or a benign ovarian mucinous cystadenoma. Particular care should be taken in cases where mucus from a benign tumor is combined with groups of mesothelial cells with reactive changes. To establish the nature of the lesion, the mucus-producing cells should be carefully examined for the presence or absence of signs of malignancy.
peritoneal fluid is a lubricating fluid (produced and absorbed by the peritoneum) found in the abdominal cavity. The abdomen is the space between the abdominal organs (such as the stomach, spleen, liver, and gallbladder) and the membrane that lines the abdominal wall.
Peritoneal fluid is a clear, sterile fluid composed primarily of water and small amounts of white blood cells, antibodies, electrolytes, and other biochemicals. The main function of peritoneal fluid is to reduce friction caused by the movement of the abdominal organs.
Reasons for analysis
In healthy people, the abdominal cavity contains a small amount of peritoneal fluid. But certain problems can lead to its excessive accumulation. This fluid, also called ascitic fluid, can lead to a condition called ascites. This is one of the complications caused by cirrhosis.
Some infections and microorganisms can also cause peritonitis, an inflammation of the peritoneal membrane.
In this case, culture of peritoneal fluid is performed. It is needed in order to diagnose the problem and begin treatment.
Culture of peritoneal fluid
This is a laboratory test in which a sample of fluid is taken from the abdominal cavity, which is then examined for microorganisms, bacteria and fungi that can cause infection.
Procedure
Some peritoneal fluid will be removed from the abdomen and sent to the laboratory for culture and gram staining. The sampling procedure is called .
Training
Before starting a laparocentesis, it is necessary to empty the bladder.
The puncture site will be cleaned with an antiseptic.
Anesthetize (using local anesthesia).
They will insert a needle (or a trakar with a canula, for which they may make a small incision) and remove a sample of fluid.
When extracting a large volume of fluid, the patient may feel slightly dizzy.
Risks associated with the procedure
There is a small risk that the needle will pierce the bladder, intestines, or blood vessel. This can lead to perforation and bleeding or infection of the intestine.
Ovarian and cervical cancer are the leading cancers leading to death in women. The insidiousness of these diseases is that they often do not manifest themselves in any way or have mild symptoms. Because of this, the tumor can grow to a serious size before it is detected. Cytological examination of the peritoneal fluid can be very helpful in detecting cancer cells or other genetic abnormalities of the ovaries and cervix at an early stage.
UDC 579.842.23+ 616-092.19
T.P. Starovoitova, T.A. Ivanova, G.B. Mukhturgin, S.A. Vityazeva, V.I. Dubrovina,
K.M. Korytov, S.V. Balakhonov
CHANGES IN THE CELLULAR COMPOSITION OF THE PERITONEAL FLUID OF WHITE MICE DURING THE INFECTIOUS PROCESS CAUSED BY YERSINIA PESTIS WITH DIFFERENT PLASMID COMPOSITION
Irkutsk Research Anti-Plague Institute of Siberia and the Far East (Irkutsk)
The article presents data on the effect of the plasmid composition of the plague microbe on the subpopulation composition of mononuclear cells in the peritoneal fluid of white mice in the early stages of the infectious process. It was shown that the change in the cellular composition of the peritoneal fluid of experimental animals depends on the plasmid profile of the plague microbe strains. During the experiment, phases were also revealed in the change in the quantitative composition of mast cells in the peritoneal fluid of white mice infected with Yersinia pestis strains with different plasmid spectrum. Keywords: Yersinia pestis, peritoneal fluid, virulence
CHANGES IN CELLULAR COMPONENTS OF PERITONEAL FLUID OF WHITE MICE WITH INFECTION CAUSED BY YERSINIA PESTIS WITH DIFFERENT PLASMID PROFILE
T.P. Starovoytova, T.A. Ivanova, G.B. Mukhturgin, S.A. Vityazeva, V.I. Dubrovina,
K.M. Korytov, S.V. Balakhonov
Irkutsk Antiplague Research Institute of Siberia and Far East, Irkutsk
The article presents the data on the influence of Yersinia pestis plasmid profile on subpopulation structure of mononuclear cells of peritoneal fluid of mice at the early stages of infectious process. It was shown that change of cellular composition of peritoneal fluid of the experimental animals depended on the plasmid profile of Yersinia pestis strains. The phase character in the changes of quantitative composition of the mast cells of peritoneal fluid of white mice infected with Y. pestis strains with different plasmid spectrum was determined. Key words: Yersinia pestis, peritoneal fluid, virulence
The vast majority of Yersinia pestis virulence factors are associated with plasmid composition. The genome of the plague pathogen of the main subspecies - Yersinia pestis subspecies pestis - has three plasmids - pYY(45MDa), pYP(6MN), and pYT(61MDa), their role in the implementation of the pathogenic properties of Yersinia has been well studied. With the presence of the pYV plasmid, Yersinia strains exhibit many phenotypic features: cell adhesion, auto-agglutination, surface agglutination, as well as the synthesis of outer membrane proteins, including V- and W-antigens and other proteins, the action of which is aimed at suppressing the phagocytic activity of immune cells. systems. Plasmids pYP and pYT are vidospecific. Plasmid pYP determines the synthesis of bacteriocin pesticin 1 and plasminogen activator, and plasmid pYT encodes two of the most well-studied virulence factors - mouse toxin and F1 capsule. A distinguishing feature of the pathogen circulating in the Tuva focus is the presence in its genome of an additional fourth plasmid pTR33 with still unclear functions. This plasmid is believed to be a genetically modified 9.5 kD resident plasmid carrying the pla (plasminogen activator) and pstl (pesticin 1) genes. The loss of plasmid leads to a change in biochemical, cultural properties, as well as to a decrease or complete loss of the virulence of the pathogen.
The leading clinical sign of plague infection and intoxication, which determines the severity of the course
and the outcome of the disease is a violation of the homeostasis of the macroorganism. Primary targets for endotoxin are polymorphonuclear leukocytes, macrophages, monocytes, endothelial cells and other cellular elements. Changes in the cellular composition of the peritoneal fluid can be regarded as a diagnostic criterion for the severity of the disease in many diseases, including plague. In this regard, the assessment of the quantitative and qualitative cellular composition of the peritoneal fluid in white mice with an infectious process caused by Y. pestis with different plasmid composition is of great interest.
Purpose of work: to study the dynamics of changes in the subpopulation composition of mononuclear cells in the peritoneal fluid of white mice in the early stages of experimental plague infection.
MATERIALS AND METHODS
The experimental model in the experiments was 175 outbred, but standard in terms of keeping conditions and weight (18-20 g) white mice of both sexes. Animals were withdrawn from the experiment in accordance with the "Rules of Laboratory Practice in the Russian Federation" approved by the Order of the Ministry of Health of the Russian Federation No. 267 dated June 19, 2003, and the National Standard of the Russian Federation GOST R 53434-2009 "Principles of Good Laboratory Practice".
We used 6 strains of Y. pestis subsp. pestis and Y. pestis subsp. altaica from the collection of the Ir-
Table 1
Characteristics of the tested strains of the plague microbe
Strain Place of isolation Plasmid composition Virulence for white mice (RbO), m.c.
Y. pestis subsp. pestis I-2638 Tuva natural plague focus pYP+pYV+pTP33+pYT+ 10 / highly virulent
Y. pestis subsp. pestis I-3479 Irkutsk Anti-Plague Institute pYP+pYV-pTP33+pYT+ avirulent
Y. pestis subsp. pestis I-3480 Irkutsk Anti-Plague Institute pYP-pYV-pTP33+pYT+ avirulent
Y. pestis subsp. altaica I-2359 Gorno-Altai natural plague focus pYP+pYV+pYT+ 4 x 104/weakly virulent
Y. pestis subsp. altaica I-2948 Gorno-Altai natural plague focus pYP-pYV+pYT+ 3 x 108/residual virulence
Y. pestis subsp. altaica I-2948/3 Irkutsk Anti-Plague Institute pYP-pYV-pYT+ avirulent
Kutsk Research Anti-Plague Institute of Rospotrebnadzor (Table 1).
Intact white mice were divided into 6 experimental and 1 control groups of 25 mice each. Animals of the experimental groups were infected with Y. pestis at a concentration of 1 x 106 mc in a volume of 0.5 ml by the intraperitoneal method. The first experimental group of animals was injected with a two-day culture of Y. pestis subsp. grown at a temperature of 28 °C. pestis I-2638, group II - Y. pestis subsp. pestis I-3479, group III - Y. pestis subsp. pestis I-3480, IV experimental group of animals were infected with the reference Gorno-Altai strain of Y pestis subsp. altaica I-2359, group V - Y. pestis subsp. altaica I-2948, group VI - with a selection strain of Y. pestis subsp. altaica I-2948/3.
Material sampling from experimental animals (peritoneal fluid) was performed after 30, 60, 90, 120 and 180 minutes. The total number of nuclear cells in 1 ml of peritoneal fluid was counted in fixed preparations stained by standard methods. For bacteriological analysis, blood from the heart and peritoneal fluid (0.1 ml each) were inoculated on a solid nutrient medium (Hottinger agar, pH 7.2).
Survey microscopy methods were used in the work. Quantitative assessment of the total number of leukocytes was carried out using a unified method for counting cells in the Goryaev chamber. The percentage of different types of leukocytes was carried out by the method of morphological examination of peritoneal fluid in smears. In the study of preparations using the computer program "MoticImagesPlus" (version 2), a differentiated count of tissue basophils (TC) was carried out, their diameter and area were measured. The degree of MC activation was assessed by the cell degranulation index (IDTC) - the percentage ratio of degranulated mast basophils to their total number.
Automatic image analysis was performed using a Zeiss light microscope (Germany) with a Moticam 2000 video camera, resolution 1392 x 1040 pixels, approx. 10, vol. one hundred.
The significance of the results of the study was obtained by mathematical methods of statistical processing using a comparative analysis according to the Student's t-test and using the computer program Statistica, version 6.0 (StatSoft Inc. 19842001, IPHI 31415926535897) and the software package
Microsoft Office Excel (2003). The results were considered significant in relation to the control at p< 0,05.
RESULTS AND DISCUSSION
The total number of cells in the peritoneal fluid in intact animals is 4.3 ± 0.9 x 103 in 1 cm3, while macrophages are the predominant cell type and account for 60.5 ± 5.6% of the total 17.0±2.8%; 5.5 ± 0.8% are mesothelial cells and other cellular elements.
In infected white mice, phasicity is observed in the change in the total number of nuclear cells. In animals infected with a virulent strain of Y. pestis subsp. pestis I-2638, after 30 min the total number of nuclear cells increases sharply to 1.5 ± 0.4 x 104 in 1 cm3, which is 3.4 times higher than in intact animals. By 60 minutes of the study, the indicators decrease to intact values, continuing to decrease in subsequent periods. The cytological picture of the peritoneal fluid has a clear relationship with the infecting culture. In animals of group I, 30 minutes after infection, an increase in the number of lymphocytes was noted, exceeding the value in intact animals by 4 times due to a sharp decrease in the number of monocytes. These changes were detected in all periods of observation. In the peritoneal fluid of white mice infected with Y. pestis subsp. pestis I-2638, after 120 minutes from the beginning of the experiment, an increase in the number of segmented neutrophils by 2.5 times was registered, compared with the control (p< 0,05), и незначительное увеличение количества палочкоядерных нейтрофилов. На последнем сроке исследования в мазках перитонеальной жидкости выявляется большое количество фибробластов, агрегация лимфоцитов и большое количество делящихся клеток.
White mice infected with Y. pestis subsp. altaica I-2359, Y. pestis subsp. pestis I-3479 and Y. pestis subsp. altaica I-2948/3, 30 min after the start of the experiment, there were no statistically significant changes. By 180 min, the number of nuclear cells in the peritoneal exudate exceeds the control values by 2.8 (p< 0,01), 1,9 (р < 0,05) и 1,5 раза соответственно. При введении животным Y. pestis subsp. pestis И-3480 и Y. pestis subsp. altaica И-2948 через 30 мин отмечается повышение общего числа ядерных клеток с последующим снижением (120 мин) до уровня контроля, и к 180 мин показатель вновь увеличивается.
When viewing smears of peritoneal fluid in animals of all experimental groups, cell proliferation of lymphocytes, histiocytes, an increase in eosinophils, tissue basophils, plasma cells, mesothelial cells and fibroblasts are recorded.
Evaluation of the morphological properties of basophils, their number and functional activity are of interest in the study of the cellular composition of the peritoneal fluid of infected animals.
It has been established that in experimental white mice there is a phase in the change in the quantitative composition of tissue basophils of the peritoneal fluid. An increase in their number in animals infected with Y. pestis subsp. pestis I-2638, is registered 60 minutes after the introduction of the culture, exceeding the values in intact animals by 2.6 times (p< 0,05). Затем данные показатели снижаются (90-120 мин) до значений ин-тактных животных, к 180 мин вновь возрастают, достигая значений 8,5 против 2,5 в контроле (р < 0,05). Часть ТК представлены интестинальными - незрелыми формами (рис. 1), появление которых можно расценивать как процесс компенсации.
Rice. 1. White mouse infected with Y. pestis subsp. pestis I-2638. peritoneal fluid. intestinal mast cells. Coloring according to Romanovsky - Giemsa, SW. x 100.
In the first terms of the study, atypical MCs accounted for 21.0 ± 1.8% of the total number of MCs, by the last terms, these figures increased to 25.2 ± 2.1%. Atypical TCs have minimal functional potential and are much smaller. The cell diameter is 6.8-8.6 μm, which is on average 2.3 times less (p< 0,05), по сравнению с диаметром типичных ТК. Таких клеток значительно меньше в перитонеальной жидкости белых мышей, зараженных Y. pestis subsp. altaica И-2359, и только в период 120-180 мин после заражения отмечаются единичные интестинальные тканевые базофилы. У животных других опытных групп атипичные ТК не выявляются.
In general, the activation of the TC system reflects the general adaptive restructuring of the body in response to the introduction of an antigen. Degranulation of tissue basophils proceeds along the path of whole-granular exocytosis (Fig. 2). The functional activity of tissue basophils in the peritoneal fluid of experimental animals has a phase character. The highest IDTK is noted
in white mice 60 minutes after infection with Y. pestis subsp. pestis I-2638 - 3.9 ± 0.6, which is 18.5 times (p< 0,01) выше значения у интактных животных, затем показатель резко снижается, но к 180 мин исследований он вновь повышается, превышая значение в контрольной группе в 4,4 раза (р < 0,01). У селекционных клонов Y. pestis subsp. pestis И-3479 и И-3480 максимальное значение индекса дегрануляции имеет место через 90 мин от начала опыта и составляет 2,0 ± 0,3 и 1,3 ± 0,4 соответственно, при этом у белых мышей II опытной группы показатели во все сроки исследования были выше, чем у животных III опытной группы.
Rice. 2. White mouse infected with Y. pestis subsp. pestis I-2638. peritoneal fluid. Mast cells. Degranulation. Coloring according to Romanovsky - Giemsa, SW. x 100.
The most pronounced phase character of changes in tissue basophils is observed in white mice of the IV experimental group. The maximum value of IDTC falls on the second and fourth stages of the study, exceeding the values of intact animals by 5.8 and 7.4 times (p< 0,05) соответственно. У особей, зараженных Y. pestis subsp. altaica И-2948/3, только на двух сроках исследования (60-90 мин) регистрируется увеличение дегрануляции тучных клеток в 3,6 и 2,6 раза соответственно (р < 0,05), в другие сроки данные статистически не значимы. У белых мышей V опытной группы максимальное значение ИДТК приходится на второй и последний срок исследования - 0,99 и 0,92 у. е., при в контроле отмечается 0,21 у. е.
CONCLUSION
Thus, the development of the infectious process in the first hours after the inoculation of the plague pathogen depends on its plasmid profile, since the most pronounced changes in the quantitative and qualitative cellular composition of the peritoneal fluid were detected in experimental animals when infected with strains containing pYP+pYV+pYT+.
The phase pattern revealed during the experiment in the change in the quantitative composition of mast cells in the peritoneal fluid, especially in individuals infected with the virulent strain of Y. pestis subsp. pestis I-2638 (pYP+pYV+pTP33+pYT+), as well as the presence of immature and atypical forms of MC, indicates the development of compensation processes.
In general, the activation of the mast cell system reflects the general adaptive restructuring of the body in response to the introduction of an antigen.
LITERATURE REFERENCES
1. Anisimov A.N. Ypestis factors that ensure circulation and preservation of the plague pathogen in ecosystems of natural foci. Message 1 // Molecular Genetics, Microbiol. and virusol. - 2002. - No. 3. - S. 3-23.
Anisimov A.N. Factors of Y. pestis providing circulation and preserving of plague infectious agent in the ecosystems of natural foci. Report I // Molekuljarnaja genetika, mikrobiol. i virusol. - 2002. - N 3. - P. 3-23. (in English)
2. Balakhonov S.V. Detection of the nucleotide sequences of the pla, pstl and cafl genes on the 33 kd cryptic plasmid of Yersiniapestis strains from the Tuva plague focus // 8th Int. Symp. on Yersinia. - Turku, Finland, 2002. - No. 10. - S. 352-355.
Balakhonov S.V. Detection of pla, pstl and cafl gene nucleotide sequences in cryptic plasmid 33 kb Yersinia pestis strains from Tuva plague focus // 8th Int. Symp. on Yersinia. - Turku, Finland, 2002. - N 10. - P. 352-355. (in English)
3. Vityazeva S.A., Starovoitova T.P., Bushkova A.V. Tissue basophils as representatives of a large cell population of the APUD system. - Dep. in VINITI No. 376-B2010 06/17/2010. - 18 s.
Vityazeva S.A., Starovoytova T.P., Bushkova A.V. Tissue basophiles as the representatives of numerous cell population of APUD-system. - Dep. in VINITI N 376-B2010 06/17/2010. - 18 p.m. (in English)
4. Krasnozhenov E.P., Fedorov Yu.V. Influence of the infectious process on the morphofunctional characteristics of tissue basophils // Zh. microbiol., epidemiol. and immunol. - 1996. - No. 1. - S. 107-108.
Krasnozhenov E.P., Fedorov Yu.V. Influence of infectious process on morphofunctional characteristics
of tissue basophiles // Zhurn. mikrobiol., jepidemiol. i immunol. - 1996. - N 1. - P. 107-108. (in English)
5. Lebedeva S.A., Trukhachev A.L., Ivanova V.S., Arutyunov Yu.I. et al. Variability of the plague pathogen and problems of its diagnosis / Ed. S.A. Lebedeva. - Rostov-on-Don: Antey, 2009. - 533 p.
Lebedeva S.A., Trukhachev A.L., Ivanova V.S., Arutyunov Yu.I. et al. Diversity of plague infectious agent and problems of its diagnostics / Ed. by S.A. Lebedeva. -Rostov-on-Don: Antei, 2009. - 533 p. (in English)
6. Menshikov V.V. et al. Laboratory research methods in the clinic. - M.: Medicine, 1987. - 365 p.
Menshikov V.V. et al. Laboratory methods of research in clinic. - Moscow: Medicina, 1987. - 365 p. (in English)
Peritoneal fluid is the substance responsible for moisturizing the abdominal wall and organs in the abdominal cavity.
It helps to prevent friction between the organs in the pelvic cavity as they move during the digestion of food.
This fluid is made up of water, electrolytes, antibodies, white blood cells, and biochemicals.
Peritoneal fluid got its name from the Latin name for the peritoneum (peritoneum), which is a serous membrane that covers the internal organs and walls of the abdominal cavity. The serous membrane is the membrane that produces fluid.
sheets of peritoneum
The peritoneum, which produces peritoneal fluid, consists of two sheets. The first sheet is the parietal peritoneum, which connects to the wall of the abdominal cavity. It is a source of fluid that moisturizes the walls of the abdominal cavity. The second sheet is the visceral peritoneum, which envelops the internal organs located in the pelvic cavity. In the visceral peritoneum, peritoneal fluid is formed, which protects the abdominal organs.Some of the abdominal organs moistened by peritoneal fluid include the liver, spleen, gallbladder, kidneys, pancreas, and stomach. Without this fluid, their movement can cause irritation in the corresponding area of the body. This can lead to infection.
While peritoneal fluid is extremely important, excess fluid can have serious consequences.
Liver disease, heart failure, and cancer of the ovaries, breast, colon, lung, stomach, and pancreas can provoke excessive production of this fluid. The term "ascites" is used to describe the presence of an excess volume of fluid in the abdominal cavity.
Symptoms of excess peritoneal fluid
Due to the severity of the diseases associated with the appearance of excess peritoneal fluid, it is very important to identify the symptoms of this condition. Some of the common symptoms include bloating, difficulty breathing, a feeling of heaviness or fullness, swelling of the legs, and blood in the vomit. For those who may have cancer, symptoms also include significant weight loss and fatigue.The first step in diagnosing the presence of excess peritoneal fluid is usually a physical examination by a doctor. If this condition is suspected, an ultrasound or CT scan may be ordered. Some more invasive procedures to confirm the presence of excess peritoneal fluid include a liver biopsy or removal of some of this fluid for analysis.