Tuberculosis remains an unconquered, dangerous, and widespread infection with the status of a socially significant disease. The unique acid‑fast envelope of Mycobacterium tuberculosis protects the pathogen against various environmental factors, the immune system, and most known chemotherapeutic agents. Moreover, the pathogen rapidly develops drug resistance, and the pathogenesis is characterized by an inherent tendency toward chronicity [1]. These factors, in accordance with chemotherapeutic principles, necessitate long‑term combination therapy using several agents with different mechanisms of action to achieve potentiation.

Based on efficacy, antituberculosis drugs are divided into three lines. In the absence of contraindications and drug resistance, the most rational choice is the combination of two first‑line agents: the antibiotic rifampicin and the synthetic hydrazide of isonicotinic acid, isoniazid. This combination remains the preferred regimen for tuberculosis chemotherapy. However, due to the rapid spread of multidrug resistance, phthisiologists are forced to switch to various combinations of second‑line reserve drugs in nearly 50% of cases [2].

In all instances of prolonged combination chemotherapy, in addition to potentiation of the antimicrobial effect, a supra‑additive enhancement of adverse effects frequently occurs. For the rifampicin‑isoniazid combination, apart from neurotoxicity, an increased damaging effect on liver function and structure is most often reported. According to some authors, the incidence of liver damage during long‑term administration of first‑line antituberculosis agents may reach 50%. Kidney dysfunction ranks second in frequency among parenchymatous organ damage during intensive antituberculosis chemotherapy [3, 4].

Since the last decade of the 20th century, cases of drug‑induced pancreatic damage have been increasingly reported. Drug‑induced pathology of this organ provoked by first‑line antituberculosis agents often manifests with symptoms mimicking other digestive disorders and is therefore not always recorded in official statistics as an adverse effect. Nevertheless, some clinicians report that drug‑induced pancreatic damage caused by first‑line drugs, presenting as chronic pancreatitis, may affect up to 22.3% of phthisiatric patients [5, 6].

Thus, the frequency of organ toxicity during tuberculosis treatment ranks liver damage first, kidney damage second, and pancreatic damage third. Combined organ damage occurs in nearly half of the cases. It should be noted that due to the specific nature of organ toxicity of the most widely used first‑line combination, the role of the primary or potentiating agent may vary depending on the organ. In particular, isoniazid exhibits primary organotoxicity toward the liver and pancreas, while nephrotoxicity is usually attributed to the damaging adverse effect of rifampicin [7].

Pharmacological protection of the structure and function of parenchymatous organs damaged by various causes, including iatrogenic factors, typically involves the use of specialised organoprotective drug classes. Hepatoprotectors are most commonly used in clinical practice, while the armamentarium for kidney and pancreas protection is very limited, partly due to the lower incidence of damage to these organs. When developing new organoprotective agents aimed at reducing the organotoxicity of antituberculosis drugs, it should be considered that prolonged antituberculosis chemotherapy is complicated by multiple adverse reactions in 80‑90% of cases, including combined parenchymatous organ damage [4]. Consequently, it is highly desirable that new organoprotectors possess the broadest possible universality, manifesting as simultaneous protective activity toward the structure and function of the liver, kidneys, and pancreas.

One of the most promising and relatively safe approaches to further improving pharmacological protection of parenchymatous organs from various toxic insults, including adverse effects of tuberculostatic agents, is the experimental identification of novel effective organoprotectors among peptide drugs [8].

Currently, in phthisiatric practice, the peptide drug glutoxim is widely used as an immunomodulator with additional hepatoprotective activity. Importantly, the hepatoprotective effect of this agent has been patented specifically for counteracting the hepatotoxicity of antituberculosis drugs [9]. Furthermore, there are reports of glutoxim’s nephroprotective activity based on clinical data indicating that this agent potentiates the therapeutic effect of fosinopril in improving renal haemodynamics in diabetic nephropathy [10].

The peptide drug dalargin is a representative of the very small group of pancreatoprotective agents. In clinical practice, it is primarily used for treating gastric and duodenal ulcers, pancreatitis, and pancreatic necrosis. In combination with octreotide, dalargin is patented for the treatment of acute pancreatitis [11]. Some researchers have suggested broader potential applications for this synthetic analogue of endogenous opioids. However, reports of successful use of this agent as a hepato‑ or nephroprotective agent could not be found in the available scientific literature.

Human chorionic gonadotropin (hCG) is likely the most promising and most universal organoprotector of peptide nature. Due to structural similarity with pituitary luteinising hormone (LH), hCG is primarily used clinically as an LH substitute for treating sexual disorders and performing IVF procedures. However, attention should be drawn to the unique ability of hCG to stimulate regeneration of various organs and tissues, which remains largely unused in clinical practice. One of the first researchers to investigate this unique activity of hCG was Professor I.M. Solopaeva from Nizhny Novgorod. Initially, under her supervision, experiments were conducted and research findings patented, demonstrating the ability of hCG to stimulate liver regeneration [12]. Subsequently, a broader concept was formulated regarding the role of hCG as a critical humoral factor stimulating growth, differentiation, and regeneration of various human organs and tissues [13]. In 2011, based on the results of in‑depth histochemical studies conducted in the USA and Canada, the ability of hCG to accelerate liver and kidney regeneration through direct stimulation of pluripotent stem cells was demonstrated and patented [14]. Over the preceding three years, members of our author team conducted priority experimental studies of the organoprotective activity of hCG and the aforementioned peptides regarding simultaneous protection of liver, kidney, and pancreatic structure and function from the toxic effects of first‑line antituberculosis drugs. Interim results of these experiments were published exclusively as abstracts [15, 16]. The completion of experimental work in this direction has enabled our team to present the results of these studies in a more comprehensive and summarised form in this article.

To conduct a comparative experimental evaluation of the hepato‑, nephro‑, and pancreatoprotective activity of three peptide drugs (glutoxim, dalargin, and human chorionic gonadotropin) using a modified model of multiorgan pathology induced by a combination of first‑line antituberculosis agents.

For experimental evaluation of organoprotective activity under conditions of first‑line antituberculosis drug administration, the liver damage model proposed by Slivka Yu.I. in 1989 is most commonly used. This model involves two‑week administration of a combination of isoniazid, rifampicin, and pyrazinamide to laboratory rats [17]. During preliminary studies attempting to precisely reproduce this model, we repeatedly encountered the near‑complete absence of morphological confirmation of liver damage in experimental rats. According to the research of Mozhokina G.N. [18], this could be explained by the ambiguous influence of sex, age, and genetic characteristics of the laboratory rats on isoniazid pharmacokinetics. Therefore, for this experiment, it was decided to enhance the damaging effect of this method of modelling toxic hepatitis with possible extension of organotoxic effects to the kidneys and pancreas. Our modification of this known pathology modelling method consisted of doubling the isoniazid dosage, extending the exposure period to 3 weeks, and replacing pyrazinamide with ethanol as a factor potentiating organotoxicity. We took into account the known facts of alcohol potentiating isoniazid toxicity [19], as well as the experience of other researchers who had previously used ethanol to potentiate the organotoxic effect of reserve antituberculosis drugs on the liver in rat experiments [20].

The subjects of this experiment were 100 male white rats weighing 230‑250 g, of which 20 animals were intact, and the remaining 80 were divided into 4 experimental groups and received intragastrically for three weeks isoniazid at a dosage of 100 mg/kg, rifampicin 125 mg/kg, and intraperitoneally 25% ethanol solution at a dosage of 3 g/kg. As experimental pharmacotherapy, animals in the first experimental group received glutoxim at a dosage of 40 mg/kg, the second group received dalargin 0.15 mg/kg, and the third group received hCG 500 IU/kg. The fourth group of experimental rats served as the control and received injections of isotonic sodium chloride solution. All drugs were administered daily subcutaneously in a volume of 0.1 ml per 100 g body weight. All animals used in the experiment were housed in the vivarium at an air temperature of 22‑24°C, with a light cycle of 12 hours light and 12 hours dark. They received standard pelleted feed and water ad libitum in compliance with all regulations and International Recommendations of the European Convention for the Protection of Vertebrate Animals Used for Experimental and Other Scientific Purposes, in accordance with the current "Good Laboratory Practice" regulations and the "Guidelines for Experimental (Preclinical) Study of New Pharmacological Substances." At the start of the experiment, all animals were healthy, with no changes in behaviour, appetite, or sleep‑wake cycles observed. Animal monitoring, including daily recording of mortality among experimental rats, was conducted over three weeks. On day 22 of the experiment, all intact rats and surviving experimental animals were euthanised by exsanguination under Zoletil veterinary anaesthesia. At the time of euthanasia, blood was collected from the carotid artery of laboratory rats for complete blood count and biochemical analysis, performed at the specialised veterinary laboratory "VetUnion" (Moscow).

Based on the results of complete blood count and biochemical analysis, the functional state of parenchymatous organs was assessed. Liver functional status was evaluated using a series of standardised biochemical blood parameters grouped by functional categories: markers of hepatocyte cytolysis (aspartate aminotransferase [AST], alanine aminotransferase [ALT], and lactate dehydrogenase [LDH] activity), indicators of synthetic activity and protein metabolism (serum cholinesterase [ChE], albumin), lipid metabolism (triglycerides, total cholesterol, low‑density lipoprotein cholesterol [LDL‑C], and high‑density lipoprotein cholesterol [HDL‑C]), liver detoxification function (direct bilirubin), and cholestasis levels (alkaline phosphatase [ALP] and gamma‑glutamyltransferase [GGT] activity, total bilirubin levels). Renal functional status was assessed by excretory function indicators (blood creatinine and urea levels) and the intensity of renal erythropoietin synthesis (red blood cell count and haemoglobin levels). The functional indicator of the endocrine portion of the pancreas was blood glucose levels, while exocrine function was assessed by blood lipase and pancreatic alpha‑amylase activity.

For morphological studies, tissue samples from the liver, kidneys, and pancreas were collected from all animals. Tissue fragments measuring 1/0.5 cm were excised from various regions of the aforementioned organs. The collected material was fixed in 10% neutral formalin, dehydrated in alcohols, and embedded in paraffin. Paraffin sections of 5‑7 μm thickness were stained with haematoxylin‑eosin, Masson’s trichrome for collagen fibres, and Hart’s fuchselin for elastic fibres. Stereometric examination of the liver, kidneys, and pancreas was performed to determine the ratios of various tissue components of these organs. Using the point‑counting method, the specific area of hepatocytes, sinusoids, and stroma was determined in the liver; the specific area of renal corpuscles, tubules, and stroma was determined in the kidneys; and acini, islets of Langerhans, and stroma were determined in the pancreas. The number of points falling on a specific tissue component was equated to its percentage content in the overall structure of the aforementioned organs, taken as 100%. Numerical data were processed using STATISTICA software (version 6) with Student’s t‑test with correction for multiple comparisons. Digital data were tested for normality of distribution. Results were considered significant if the standard error did not exceed 5% (p < 0.05).

The applied method of modelling the studied pathology using elevated dosages of first‑line antituberculosis drugs combined with alcohol proved to be highly toxic and was accompanied by 70% mortality in the control group. Administration of the studied drugs significantly improved the survival of experimental rats, indicating the presence of sufficiently pronounced antitoxic activity in these agents. Specifically, dalargin administration increased survival 1.7‑fold compared to controls, while glutoxim and hCG administration increased this parameter 2.2‑ and 2.3‑fold, respectively (Fig. 1).

Fig. 1. Effect of experimental pharmacotherapy on survival of experimental rats during 3‑week intoxication with tuberculostatics combined with alcohol.

According to biochemical blood analysis results, three‑week exposure to the combination of isoniazid, rifampicin, and ethanol provoked serious functional disturbances in all three studied organs in experimental rats, with the liver expectedly suffering most severely. Specifically, analysis of the dynamics of cytolytic enzyme activities in blood showed that against the background of modelled pathology, there was a significant increase in AST, ALT, and LDH activities by 148%, 66%, and 172%, respectively (Table 1). Only glutoxim completely prevented the increase in cytolytic enzyme activities in blood, while the therapeutic efficacy of hCG and dalargin was less pronounced.

Table 1. Effect of experimental pharmacotherapy on the activity of indicator cytolytic enzymes

Experimental GroupALT (U/L)AST (U/L)LDH (U/L)Intact66.3±5.6139.4±9.81065.6±121.6Control189.4±14.3*231.5±20.1*2901.8±196.9*Glutoxim87.5±7.1**115.5±12.7**1384.8±106.7**Dalargin112.5±14.0*/**178.6±16.8**1802.4±177.6*/**hCG108.0±12.3*/**156.2±13.3**1663.5±150.2*/**

Notes: * — significant difference compared to intact (p < 0.05); ** — significant difference compared to control (p < 0.05).

It is important to note that albumin and serum cholinesterase (ChE) synthesis occurs exclusively in the liver. In control rats, 21‑day modelled pathology caused a significant decrease in blood albumin levels by 46% and ChE activity by 37%, clearly indicating impaired protein synthesis in hepatocytes (Table 2). Among the tested drugs, only glutoxim administration normalised some of the disturbed parameters.

Table 2. Effect of the studied drugs on the protein‑synthetic function of the liver

Experimental GroupChE Activity (U/L)Albumin Content (g/L)Intact330.4±35.135.50±3.11Control178.6±19.3*22.20±2.44*Glutoxim357.5±37.1**24.83±3.02*Dalargin198.1±18.3*25.12±2.20*hCG201.6±21.8*26.04±3.10*

Notes: * — significant difference compared to intact (p < 0.05); ** — significant difference compared to control (p < 0.05).

The modelled intoxication caused an increase in ALP and GGT activity in control animals by 3.86 and 7.05 times compared to intact rats, indicating the provocation of pronounced cholestasis (Table 3). Furthermore, a sharp increase in total and direct bilirubin concentrations by 10.21 and 15.08 times was recorded, confirming the development of cholestasis and indicating intense inflammatory processes and hepatocyte death. Course administration of the studied drugs significantly reduced inflammation intensity, with hCG and glutoxim being the most effective.

Table 3. Effect of the studied drugs on changes in blood serum parameters reflecting the intensity of cholestasis and inflammatory damage to liver cells

Experimental GroupALP (U/L)GGT (U/L)Total Bilirubin (μmol/L)Direct Bilirubin (μmol/L)Intact7.64±0.632.1±0.22.14±0.221.20±0.12Control29.45±1.46*14.8±0.9*21.84±1.96*18.1±1.91*Glutoxim22.67±1.82*/**13.5±0.9*6.32±0.62*/**3.47±0.42*/**Dalargin27.08±1.38*12.6±1.0*9.41±1.60*/**6.30±0.60*/**hCG22.52±1.44*/**10.2±0.9*/**6.85±1.02*/**4.22±0.51*/**

Notes: * — significant difference compared to intact (p < 0.05); ** — significant difference compared to control (p < 0.05).

According to complete blood count and biochemical analysis results, in addition to the liver, the modelled pathology provoked pronounced renal functional disturbances in laboratory rats. In control animals, this manifested as significant increases in blood urea and creatinine concentrations by 100% and 55%, respectively, indicating impaired renal excretory function and inflammation in this organ (Table 4).

Table 4. Effect of the studied drugs on the indicators of the functional state of the kidneys

Experimental GroupUrea (mmol/L)Creatinine (μmol/L)Erythrocytes (10¹²/L)Haemoglobin (g/L)Intact7.71±0.8853.15±3.457.18±0.62138.5±8.80Control15.43±1.14*82.22±6.05*4.14±0.80*88.6±9.11*Glutoxim10.72±0.92**64.72±3.96**5.21±0.56*101.0±15.45*Dalargin11.45±1.06*/**73.84±5.20*5.60±0.52*106.6±10.7*hCG8.64±0.76**57.58±5.81**6.19±0.59**128.5±10.65**

Notes: * — significant difference compared to intact (p < 0.05); ** — significant difference compared to control (p < 0.05).

Furthermore, intoxication with tuberculostatics combined with alcohol exposure caused impaired renal erythropoietin synthesis, manifested in the complete blood count as decreased erythrocyte count and haemoglobin concentration, which decreased in control animals by 42% and 36%, respectively. Only hCG demonstrated the ability to effectively normalise red blood cell parameters dependent on erythropoietin levels. This same drug most effectively normalised urea and creatinine levels, although glutoxim and, to some extent, dalargin also caused significant positive changes in these indicators characterising renal excretory function.

The modelled pathology also negatively affected pancreatic function. Both the endocrine and exocrine portions of this organ were affected, expressed in control animals as a 66% increase in blood glucose and a 24% and 54% suppression of amylase and lipase activities, respectively (Table 5). hCG and dalargin demonstrated significant organoprotective activity regarding pancreatic function protection, with dalargin showing more pronounced efficacy, although lipase activity was not restored by dalargin, consistent with data from some clinicians who investigated the cytoprotective effects of this drug in treating post‑gastroresection pancreatitis [21].

Table 5. Effect of the studied drugs on the functional state of the pancreas

Experimental GroupGlucose (mmol/L)Pancreatic Alpha‑Amylase (U/L)Lipase (U/L)Intact5.75±0.401718.8±56.0208.0±15.6Control9.55±0.86*1312.4±51.0*94.8±7.6*Glutoxim8.25±1.04*1343.2±68.3*122.0±10.9*Dalargin5.93±0.67**1591.1±77.8**104.2±17.1*hCG6.78±0.70**1535.2±64.4**180.0±20.3**

Notes: * — significant difference compared to intact (p < 0.05); ** — significant difference compared to control (p < 0.05).

Histological examination of liver tissue from control rats showed that the modelled pathology provoked severe alternative and inflammatory changes in this organ, manifested as disorganisation of hepatic lobules, diffuse hydropic and balloon protein degeneration, as well as large‑droplet fatty degeneration of hepatocytes (Fig. 2, A). In the enlarged portal tracts, pronounced lymphocytic inflammatory infiltration was observed, but without extension into the lobules, indicating the presence of low‑grade interstitial hepatitis. Dalargin treatment led to partial preservation of liver histoarchitecture, although in some lobules, trabecular disorganisation was still observed. Hepatocytes showed focal protein hydropic degeneration (Fig. 2, B), and stromal manifestations of interstitial hepatitis were present as in the control series. Glutoxim pharmacotherapy led to normalisation of organ structure with preservation of lobular architecture. In some hepatocytes, hydropic protein degeneration persisted, but the vast majority of cells showed no degenerative changes (Fig. 2, C). In the portal tracts, not diffuse but scattered, sparse lymphocytic infiltrates were observed, i.e., interstitial inflammation was significantly less pronounced compared to the control series. Administration of hCG as pharmacotherapy led to complete restoration of liver structure without degenerative changes (Fig. 2, D). A significant portion of portal stroma showed no inflammatory infiltration, with only occasional small lymphocytic aggregates, i.e., interstitial inflammation was present to a minimal degree compared to the control series.

Fig. 2. Structural changes in the liver in the control (A), after treatment with dalargin (B), glutoxim (C), and hCG (D)

Haematoxylin and eosin (A, B, C) and Masson's (D) staining using brilliant green (×200 — A, C, D; ×160 — D). A — large‑droplet fatty degeneration of hepatocytes. B — focal protein hydropic degeneration of hepatocytes. C — normal liver structure, no degenerative changes in hepatocytes. D — portal tract with congestion of the portal vein branch, without inflammatory infiltration; hepatocytes without degenerative changes.

Stereometric examination of the liver in the control series showed (Fig. 3) that the specific area occupied by hepatocytes decreased 1.2‑fold compared to the intact series (p=0.05), while the area of sinusoids and stroma increased 1.1‑fold (p=0.05), quantitatively reflecting the development of damage and interstitial inflammation. With dalargin treatment, the specific area of hepatocytes, sinusoids, and stroma showed no significant changes compared to the control series. Stereometry in the glutoxim series established that the specific area of hepatocytes increased 1.1‑fold compared to the control series (p=0.05), sinusoid areas showed no significant changes, and stromal area decreased 1.4‑fold (p=0.001), objectively reflecting reduced inflammatory response. Stereometric examination in the hCG series showed that the specific area of hepatocytes increased 1.1‑fold compared to the control series (p=0.05), sinusoid areas showed no significant changes, and stromal area decreased 1.4‑fold (p=0.001), similar to the glutoxim series.

Fig. 3. Effect of experimental pharmacotherapy on liver tissue stereometry

Notes: * — significant difference compared to intact (p < 0.05); ** — significant difference compared to control (p < 0.05).

Histological examination of kidneys from control animals revealed epithelial cells of proximal and distal convoluted tubules in a state of protein hydropic, hyaline‑droplet degeneration, and coagulation necrosis compared to the intact group (Fig. 4, A). Tubular lumens were markedly dilated and filled with eosinophilic proteinaceous and necrotic material (Fig. 4, A), hyaline casts, and basophilic calcium deposits with aggregates of segmented neutrophils. Additionally, glomerulosclerosis and stromal sclerosis were detected in renal tissue, with diffuse lymphocytic inflammatory infiltration (interstitial nephritis). Glomeruli appeared normal. With dalargin use, the renal tubular apparatus showed structural changes comparable to the control series. This was expressed as epithelial cells of the main nephron segments exhibiting hyaline‑droplet and hydropic degeneration with necrosis of individual cells (Fig. 4, B). The renal stroma of both cortical and medullary layers showed preserved interstitial inflammation. Glomeruli remained unchanged. With glutoxim treatment, epithelial cells of proximal and distal convoluted tubules showed hydropic degeneration with cytoplasmic vacuolisation (Fig. 4, C). In this series, no hyaline‑droplet degeneration or necrosis was detected. Focal lymphocytic infiltrates were found in the renal stroma, but no diffuse infiltration was observed. hCG treatment led to near‑complete normalisation of renal structure. Epithelial cells of proximal and distal convoluted tubules showed no hyaline‑droplet degeneration or signs of coagulation necrosis. However, hydropic degeneration was still observed in some epithelial cells (Fig. 4, D). Rare, small lymphocytic aggregates were found in the renal stroma.

Fig. 4. Structural changes in the kidneys in the control (A), after treatment with dalargin (B), glutoxim (C), and hCG (D)

Haematoxylin and eosin staining (×200 — A, B, D; ×160 — C). A — hydropic and hyaline‑droplet degeneration of convoluted tubular epithelium with transition to necrosis (karyolysis), tubular lumens filled with proteinaceous eosinophilic material; glomerulus unchanged. B — hydropic and hyaline‑droplet degeneration of convoluted tubular epithelium with transition to necrosis (karyolysis), tubular lumens filled with proteinaceous eosinophilic material. C, D — hydropic degeneration of individual convoluted tubular epithelial cells, glomeruli unchanged.

Stereometric examination of renal tissue in the control group showed that the specific area of renal corpuscles decreased 1.4‑fold compared to intact animals (p=0.001), while the area occupied by tubules showed no significant changes (Fig. 5). Stromal area increased 1.3‑fold (p=0.001), objectively confirming the development of pronounced interstitial inflammation and sclerosis. Dalargin treatment was accompanied by no significant changes in the specific area of renal tissue structures, similar to the liver findings. Stereometric examination in the glutoxim group showed that the specific area of renal corpuscles and tubules showed no significant changes, while stromal area decreased 1.2‑fold (p=0.05). In the hCG group, the specific area of renal corpuscles increased 1.3‑fold (p=0.001), tubular area decreased but not significantly, while stromal area decreased 1.3‑fold (p=0.001).

Fig. 5. Effect of experimental pharmacotherapy on renal tissue stereometry

Notes: * — significant difference compared to intact (p < 0.05); ** — significant difference compared to control (p < 0.05).

Histological examination of the pancreas from control rats revealed that most islets of Langerhans underwent atrophy and sclerosis, while some islets were hypertrophied. Focal necrotic changes in acini were observed, involving interlobular and peripancreatic tissue (Fig. 6, A). With dalargin pharmacotherapy, in contrast to the histological findings in the liver and kidneys, the pancreas showed structural normalisation. No destructive changes in acini or reactive inflammation were observed in the lobules. The islets of Langerhans varied in shape and size, with segmental sclerosis of interlobular stroma preserved (Fig. 6, B). The interlobular and intralobular stroma showed infiltration by single lymphocytes, indicating a significant reduction in the degree of interstitial inflammation in the organ. With glutoxim use, pancreatic structural changes in experimental animals did not differ from controls. Specifically, islets of Langerhans decreased in size, acinar lumens were dilated and filled with secretion. Necrotic changes were noted in the lobules (Fig. 6, C). The intralobular and interlobular stroma of the pancreas was enlarged, diffusely infiltrated by lymphocytes and segmented leukocytes, and showed proliferation of coarse fibrous connective tissue. hCG treatment in rat pancreas was accompanied by normal islet of Langerhans size, absence of necrotic changes in lobules, and normal acinar lumens. The intralobular and interlobular stroma was slightly enlarged and moderately infiltrated by lymphocytes (Fig. 6, D).

Fig. 6. Structural changes in the pancreas in the control (A), with the use of dalargin (B), glutoxim (C), and hCG (D)

Haematoxylin and eosin staining (×160 — A, B, C; ×200 — D). A — necrosis of lobules with acini and interlobular stroma. B — moderate lymphocytic infiltration of interlobular stroma and sclerosis. C — necrosis of lobules with acini and interlobular stroma. D — normal glandular structure with islets of Langerhans and unchanged acini.

Stereometric results of pancreatic tissue structures (Fig. 7) in the control series established that the specific area of islets of Langerhans decreased 1.3‑fold (p=0.05), while stromal area increased 1.3‑fold (p=0.05), reflecting the development of inflammatory response in the interstitium. The specific area of acini showed no significant changes compared to intact animals. In the dalargin group, the specific area of islets of Langerhans increased 1.7‑fold (p=0.001), while stromal area decreased 1.5‑fold (p=0.001), reflecting reduced inflammatory response and restoration of organ structure. The specific area of acini showed no significant changes. Glutoxim treatment resulted in no significant changes in the specific area of islets of Langerhans and acinar apparatus, while stromal area decreased 1.2‑fold (p=0.05) compared to the control series. In the hCG group, the specific area of islets of Langerhans increased 1.4‑fold (p=0.001), stromal area decreased 1.2‑fold (p=0.05), while acinar area showed no significant changes compared to the control series.

Fig. 7. Effect of experimental pharmacotherapy on pancreatic tissue stereometry parameters

Note: * — significant difference compared to intact (p < 0.05).

The present study established that the applied experimental model of three‑week intoxication in laboratory rats with a combination of first‑line antituberculosis agents (isoniazid 100 mg/kg and rifampicin 125 mg/kg) with additional daily intraperitoneal injections of 25% ethanol solution as an organotoxicity‑potentiating factor at a dosage of 3 g/kg caused severe intoxication in experimental animals, resulting in mortality of 70% of untreated (control) animals. In surviving laboratory animals at the end of the experiment, biochemical blood analysis and in‑depth histological studies revealed the development of toxic multiorgan pathology accompanied by severe structural and functional disturbances in three parenchymatous organs: the liver, kidneys, and pancreas, due to a pronounced chronic inflammatory process. The applied model of multiorgan pathology can fundamentally be used in preclinical studies for testing new organoprotective agents, whose efficacy should be evaluated by preventing mortality and the degree of reversibility of pathological shifts in both indicator biochemical parameters and structural changes in the mentioned organs based on histological findings. To reduce the number of laboratory animals used in the future, it would likely be advisable to reduce the mortality rate in controls to 40‑50% by lowering the ethanol dosage and changing the route of administration from intraperitoneal to intragastric.

Testing of this multiorgan pathology model provided experimental evidence supporting the feasibility of expanding the indications for glutoxim and dalargin for use as organoprotective agents during combination tuberculosis treatment. Furthermore, a set of experimental data was obtained substantiating the promising prospects for initiating preclinical trials of hCG for the additional indication as an effective and sufficiently universal organoprotective agent.