Cardioprotective, anti-inflammatory, and antioxidative outcome of costus against bleomycin-induced cardiotoxicity in rat model

Abstract

Due to bleomycin’s cytotoxic characteristics, which include cardiotoxicity, this investigation looked at the effectiveness of costus ethanolic extract in reducing cardiotoxicity in male rats receiving bleomycin therapy. Forty adult male rats (160–200 g) were evenly allocated into four groups: group (1) included normal rats serving as the control; group (2) included normal rats administered 200 mg/kg of costus ethanolic extract (CEE) orally for 6 weeks; group (3) consisted of rats receiving bleomycin (15 mg/kg twice weekly, ip) for 6 weeks; and group (4) involved rats treated orally with CEE (200 mg/kg/day) for 6 weeks following bleomycin intoxication. The results indicated that the CEE significantly reversed the cardiological deteriorations brought on by bleomycin; this was demonstrated by a considerable increase in cardiac SOD, GPx, GSH, and CAT, along with a substantial decrease in cardiac MDA, NO, and DNA fragmentation. Also, serum, LDH, CK-MB, CK- total, TNF-α, IL-4, IL-6 IL-10, IL-1β, triglycerides, cholesterol, and LDL were significantly reduced, while CD4 levels increased, and HDL declined significantly. The results of the histological and immunohistochemical analyses revealed a notable regeneration. In conclusion, CEE’s anti-cardiotoxic, anti-inflammatory, and antioxidant properties prove its ability to be a cardio-protective supplement. This may be mediated by its active constituents’ radical scavenging and antioxidant properties, particularly high phenolic content.

1. Introduction

Cancer was the second most prevalent cause of death in 2018.¹ Along with radiotherapy and surgery, chemotherapy is a crucial cancer treatment. But its cardiotoxicity, which causes cardiomyopathy, is still a serious side effect.² Heart failure results from cardiomyopathy, which causes the heart's pumping capacity to decline. It is the third contributing cause of heart failure after coronary insufficiency and hypertension.3, 4 Many types of cancer, such as Hodgkin's reticulosarcomas, malignant non-lymphomas, and certain reproductive cancers, can anticipate living longer and having a good quality of life appreciation to the widespread usage of various chemotherapy regimens, which improves their prognosis. The oncological pathology mentioned above is frequently treated with bleomycin.⁵ Bleomycin, an antibiotic that fights tumors, was discovered as an A2 fraction in a culture of Streptomyces verticillus. Its toxic effects on lung tissue, including the onset of pulmonary fibrosis, pleurisy with pain, and worsening respiratory failure, are among the first-known side effects. Infrequent toxic effects on blood vessels encompass cerebral arteritis, stroke, myocardial infarction, thrombotic microangiopathy, and Raynaud's syndrome.⁶ It may harm blood vessels, which may occasionally result in Raynaud's syndrome, cerebral arteritis, stroke, myocardial infarction, and thrombotic microangiopathy.⁵ The effects of medicine on the myocardium, particularly when administered intravenously, are also unknown. Cardiotoxicity is the most frequent symptom of treatment-related morbidity, even though modern advancements in cancer treatments, such as chemotherapy and radiation therapy, have boosted the survival rate of cancer patients.⁷ Cardiotoxicity refers to the complications that include various adverse effects that lead to cardiac dysfunction on the background of the drug and radiotherapy⁸. Bleomycin (BLM) is a medication produced by S. verticillus bacteria and is classified as a glycoprotein antibiotic. It is an antitumor drug used to treat malignancies such as lymphomas, testicular carcinoma, squamous cell carcinoma, and malignant pleural effusion.⁹ Among the frequent side effects of BLM medication include thrombotic microangiopathy, cerebral arteritis, myocardial infarction, pulmonary fibrosis, and Raynaud's syndrome.¹⁰ Reports indicate that the myocardium exhibits atrophic and inflammatory alterations following a single BLM injection.¹¹.

There is currently no evidence connecting the acute chest pain brought on by bleomycin infusion to a pathophysiological cause. One likely reason is grave inflammation, which might manifest as acute pleuro-pericarditis or more widespread mucocutaneous toxicity related to bleomycin therapy. It is crucial to consider the vascular etiology of the pain because several pulmonary vascular illnesses, including pulmonary hypertension and pulmonary embolism, can result in pleuritic and substernal chest pain, even in the loss of infarction. Regrettably, there is still uncertainty about the anatomical mechanism behind the cardiotoxic effects of bleomycin. Due to their beneficial chemical components, herbal medicines have been utilized to treat numerous diseases for over 4,000 years. Essentially, the phytochemical components of plants that carry out precise pharmacological effects when consumed by humans are where their medicinal potential lies.12, 13.

The plant Saussurea costus, also known as Saussurea lappa Clarke,¹⁴ is a constituent of the Asteraceae family and is referred to as the Kuth root or Indian costus.15, 16 Costus was used as a medicinal plant to cure various conditions, including asthma, inflammatory diseases, ulcers, and stomach issues. It was utilized in modern medicine and was listed in the Prophet's medicine to treat numerous diseases.16, 17 A plant with a high concentration of antioxidants is Saussurea costus.¹⁸ Its pharmacological effects include anti-inflammatory, immune-modulating, hypoglycemic, anti-hepatotoxic, hypolipidemic, antiparasitic, antiviral, and anticancer properties.19, 20 Therefore, the current investigation aimed to evaluate the cardioprotective efficacy of CEE administration in bleomycin-induced cardiac damage in animal models.

2. Materials and methods

2.1. Chemicals

Bleomycin was sourced from Sigma Aldrich, located in St. Louis, MO, USA.

2.2. Plant materials and extraction

Scientific botanists identified and verified the costus roots before purchasing them from Imtinan Company in Nasr City, Cairo, Egypt. The plant was found to have a taxonomic serial number of 780691. According to the modified approach of²¹, the ethanolic extract of the dry powdered roots was performed using the previously published technique, and the 1,1-diphenyl-2-picrylhydrazyl radical scavenging activity of CEE was assessed²¹. The extract's reducing power was evaluated using the method outlined.²²The total phenolic content (TPC) of CEE was determind, using established biochemical method ⁵⁹. The total extract yield was calculated according to ⁶⁰

2.3. HPLC analysis of phenolic constituents

Using an Agilent 1260 series, the HPLC (high-performance liquid chromatography) test was performed for CEE screening. A Kromasil C18 column with a 4.6 mm ID and a 250 mm ID was used for the separation (5 m). Water and acetonitrile were the main components of the mobile phase, which had a flow rate of 1 ml/min and a trifluoroacetic acid concentration of 0.05 % (A and B). The mobile phase was programmed using a linear gradient at the following intervals: 0 min (82 % A), 0 min to 5 min (80 % A), 5–8 min to 6 min, 5–8 min to 12 min, 85 % A, and 15–16 min (82 % A). At a wavelength of 280 nm, the multiwavelength detector was observed. Then, all the sample solutions were exposed to an injection of adequate volume (10 ml). A constant 35 °C was kept in the column temperature.

2.4. Animals and the Design of Experiments

At the National Research Centre in Giza, Egypt's Animal Colony provided male albino rats (weighing between 160 and 200 g). After acclimatization to the experimental conditions, the animals were divided into four groups of ten. In the first group, designated as a control group, healthy animals received standard food and an intraperitoneal injection of 1 ml isotonic saline. The second group consisted of good-health animals that received 200 mg/kg/day of CEE (costus ethanolic extract) orally for six weeks.²³ The third group included animals receiving 15 mg/kg of bleomycin (IP) twice a week.²⁴ The fourth group served as positive control, which included rats administered CEE orally once daily for six weeks following bleomycin intoxication.

2.5. Samples of blood and tissue

The rats underwent overnight fasting and weighing after the treatment period (6 weeks). After the intramuscular administration of sodium pentobarbital (9.1 mg/kg in a sterile 0.9 % NaCl solution), blood samples were collected from the retro-orbital plexus utilizing heparinized and sterilized glass capillaries. The blood samples underwent centrifugation for 10 min at 1000 rpm under cooling conditions to facilitate the separation of sera. The sera were subsequently divided into aliquots and stored at −80 °C. After blood collection, the animals were euthanized, and each heart was examined. For biochemical analysis, one portion of the heart was cleaned in saline, dried, and wrapped in aluminum foil. Formalin-saline (10 %) buffer was used to soak a second section of the heart for histological and immunohistochemical processing and microscopic examination.

2.6. Biochemical determinations

All biochemical measurements were performed using a Shimadzu spectrophotometer (UV–vis 1201, Japan). The serum lipid profile was determined using kits obtained from DiaSys Diagnostic Systems GmbH in Germany. Reagent kits from BioVision, based in South Milpitas, California, USA, were utilized to conduct colorimetric assays for LDH, CK-total, and CK-MB.

2.7. Markers of oxidative stress in cardiac tissue

To assess the cardiac stress, the SOD, GPx, GSH, CAT, NO, and MDA activities in cardiac rat tissues were determined based on the instructions in the enclosed guides of the commercial kits (My BioSource, San Diego, USA).

2.8. Determination of pro-inflammatory cytokine and apoptotic biomarker

Commercial rat ELISA kits were procured from MyBioSource Company (San Diego, USA). The pro-inflammatory cytokine and apoptotic biomarkers comprising IL-4, IL-6, IL-10, TNF-α, IL-1β, and CD4 were evaluated by ELISA Kits using a commercially existing highly sensitive ELISA kit to the manufacturer’s directions following the methods.25, 26.

2.9. Cardiac DNA fragmentation %

The DNA fragmentation in cardiac tissues was assessed using ELISA Kits using the quantitative method.²⁷.

2.10. Histopathology assay

After the experiment, heart samples were removed and fixed in neutral buffered formalin (10 %). Before being divided into tiny pieces (5 m), the samples were washed in xylene, embedded in paraffin, and then gradually dehydrated in ethyl alcohol. Following deparaffinization, samples were stained with Hematoxylin and Eosin under 100X light microscope examination.²²

2.11. Immunohistochemistry study

The immunohistochemical procedure was hired to determine the immunoexpressing of Caspase 3 in cardiac tissues by utilizing the rabbit polyclonal antibody CASPASE 3 as described by28, 29. The immunostaining intensity of anti-Caspase 3 in cardiac rat tissues was assessed using the Image J program.28, 29.

2.12. Statistical analysis

A post hoc (Tukey) multiple comparisons test at p > 0.05 was employed after a one-way analysis of variance (ANOVA) to compare means. This was accomplished using the statistical analysis system (S.A.S.) computer software; Copyright (c) 1998 by S.A.S. Institute Inc., Cary, North Carolina, U.S.A.

3. Results

3.1. HPLC analysis of phenolic constituents

The statistics below demonstrate the radical scavenging activity (RSA), yield, total phenolic content, and reduction of the power of the CEE (Fig. 1, Fig. 2). Most of the 16 phenolic compounds were discovered using HPLC analysis in CEE. Among the chemicals found were high amounts of naringenin, taxifolin, ferulic acid, gallic acid, chlorogenic acid, and coffeic acid (Table 1 and Fig. 3).

Fig. 1.

The yield percentage of total phenolic content and radical scavenging activity percentage of three replicates of ethanolic extract from costus dry powdered roots.

Fig. 2.

Assessment of the reducing power of three replicates of the ethanolic extract from powdered dry roots of Costus.

Table 1.

The primary phenolic compounds of the CEE (ethanolic extract of costus) were identified using HPLC analysis.

Constituents	Area	Concentration (µg/ml = µg/ 6.8 mg)	Concentration (µg/g)
Naringenin	654.73	40.05	1494.47
Chlorogenic acid	508.86	39.58	1477.00
Gallic acid	77.66	6.23	232.49
Ferulic acid	537.79	16.74	624.59
Taxifolin	82.11	9.36	349.28
Coffeic acid	87.66	3.17	118.27
Pyro catechol	28.89	2.80	104.38
Coumaric acid	150.73	2.43	90.79
Vanillin	143.40	2.24	83.58
Syringic acid	63.85	2.17	81.15
Kaempferol	20.59	1.68	62.50
Methyl gallate	4.14	0.06	2.33
Cinnamic acid	70.85	0.75	27.91
Ellagic acid	6.58	0.38	14.08
Rutin	0.00	0.00	0.00
Catechin	0.00	0.00	0.00

Fig. 3.

The results of HPLC screening of phenolic ingredients of CEE (costus ethanolic extract).

3.2. Biochemical determinations and oxidative stress markers of cardiac tissue

Contrary to predictions, bleomycin intoxication caused a significantly higher level of triglycerides, LDL, and cholesterol in the blood and a significantly inferior level of HDL related to the standard treatment. Administering CEE to bleomycin-intoxicated rats resulted in a discernible enhancement in the lipid profile as indicated by the significant reductions in LDL, triglycerides, and cholesterol, as well as the observable augments in the levels of HDL when compared with the bleomycin-treated rats (Table 2).

Table 2.

Effects of Bleomycin and/or CEE oral administration on serum lipid profile of adult male Wistar.

	Control	CEE1	Bleomycin	Bleomycin ∼ CEE
Cholesterol (mg/dl)	130.3±4.6c	128.3±6.6c	320±13.5a	204±3.9b
Triglycerides (mg/dl)	155±8.6c	152±7.7c	364±20.1 a	215±8.3b
HDL-C (mg/dl)	46.5±5.4c	45.6±4.7c	28.5±2.9 a	36.7±4.0b
LDL-C (mg/dl)	84.5±5.5c	83.7±4.9c	196.5 ± 12.5 a	110.5±6.4b

¹CEE; costus ethanolic extract. The values are stated as the mean ± SE.

^{a, b, c} Rows have significantly diverse superscripts (a, b, and c).

The significant deterioration of cardiac oxidative stress induced by bleomycin toxicity was shown by a marked increase in cardiac MDA and NO levels, with a considerable reduction in the activities of CAT, SOD, and GPx, as well as GSH concentrations (Table 3). Rats that were given CEE displayed a significant decrease in cardiac MDA and NO levels compared to the bleomycin-intoxicated group and a noticeably increased level of GSH, CAT, SOD, and GPx activities.

Table 3.

Effect of Bleomycin and/or CEE oral administration on cardiac oxidative stress.

	Control	CEE1	Bleomycin	Bleomycin ∼ CEE
MDA (pg/mL)	213.5±16.5c	210.1 ± 18.1c	487.4±40.3 a	263.7±25.3b
NO (µmol/L)	25.1±1.4c	24.4±2.2c	62.5±3.1 a	39.3±2.8b
GSH (ng/mL)	66.2±4.3c	70.3±2.8c	24.11±2.3 a	52.08±3.4b
SOD (U/L)	35.2±0.1c	38.4±1.3c	14.4±1.0 a	28.3±1.2b
GPx (U/L)	762±47c	789±37c	369±27.3 a	599±51.3b
CAT (U/L)	11.9±0.8c	12.7±0.4c	5.4±0.2 a	9.8±3.1b

¹CEE; costus ethanolic extract. The values are stated as the mean ± SE.

^{a, b, c} Rows have significantly diverse superscripts (a, b, and c).

3.3. Determination of pro-inflammatory cytokine, apoptotic biomarker, cardiac enzymes, and cardiac DNA fragmentation %

The data collected showed a significant increase in IL-10, IL-6, TNF-α, IL-4, IL-1β, LDH, CK-total, and CK-MB and DNA damage in the bleomycin group as related to the controlled one. Intriguingly, the administration of CEE to bleomycin-treated rats improved all inflammatory cytokines, apoptotic markers, DNA damage, and cardiac enzyme to within normal values while significantly lowering levels of TNF-α, IL1β, IL-4-, IL-6, IL-10, DNA damage LDH, CK-MB, and CK-total and increasing CD4 compared to bleomycin (Fig. 4a-j).

Fig. 4.

(a – j). Levels of TNF-α, IL-1β, IL-4, IL-6, IL-10, CD4, CK-MB, CK-total, and LDH, together with cardiac DNA fragmentation in control, bleomycin-intoxicated, and CEE-treated male rats. * Exhibits a significant difference from the control group, while # demonstrates a significant difference from the bleomycin group (p ≤ 0.05).

3.4. Histopathology

Examining the histological architecture of H&E-stained tissues of the control group revealed normal myocardial fiber. The histological architecture of H&E-stained tissues of the control group revealed normal myocardial fiber. The histological architecture of H&E-stained tissues of the control group revealed normal myocardial fiber arrangement with typical striation and branching of muscle fibers. The cardiomyocytes exhibited central oval nuclei and acidophilic sarcoplasm. The intercellular spaces infiltrated with few blood capillaries (Fig. 5a). CEE–treated group showed approximately typical cardiac muscle structure (Fig. 5b). Histological examination of the bleomycin-intoxicated group revealed marked distortion, fragmentation, signs of vacuolar degeneration in the form of hyperacid philia and cytoplasmic vacuolation, dull striations following the necrosis of myofibrillar tissue and degenerated myofibrils lost their nuclei as compared to the control group (Fig. 5c). Bleomycin-intoxicated and CEE–treated group showed marked improvement and preservation of intercellular spaces and morphology of cardiomyocytes compared to the bleomycin-treated group. Normal cardiac muscle cells with minimal cytoplasmic vacuolization were observed (Fig. 5-d).

Fig. 5.

(a-d). Histopathological examination of cardiac tissues of the groups. (a) Photomicrograph of the cardiac tissue of the control group showing the normal histological pattern of myocardial fibers (arrow) (b) Photomicrograph of CEE-treated group showing the cross-section of preserved cardiomyocyte morphology with central oval vesicular nuclei, exhibiting approximately normal histological architecture. (c) Photomicrograph of the bleomycin-intoxicated group showing degenerated cardiomyocytes (arrow). Also, vacuolization within myocardial fibers (arrowhead) indicated vacuolar degeneration in cardiomyocytes. The vacuoles could represent cardiac muscle fiber auto-phagocytosis. (d) Photomicrograph of bleomycin-intoxicated and CEE–treated group showing reduced perivascular fibrosis, with approximately normal pattern of myocardial fibers, with some residual fibers with enlarged nuclei (arrow).

3.5. Immunohistochemistry

The Caspase-3 expression was modest in control of animals. At the same time, it was moderate in the CEE group (Fig. 6a, b). Rats exposed to bleomycin displayed highly expressed caspase-3 (Fig. 6c). A modest Caspase-3 positivity was seen in the bleomycin-intoxicated and CEE-treated group, which was less immunoreactive than the bleomycin group (Fig. 6d; Fig. 7)

Fig. 6.

(a-d). Photomicrographs of cardiac tissues of various groups immune-stained with Caspase-3 (x400). (a): rats in the control groups have normal weak cytoplasmic Caspase-3 IHC positivity. (b): Rats treated with CEE display mild cytoplasmic Caspase-3 activity. (c): Rats from the bleomycin-intoxicated group display a severe Caspase-3 activity (arrow). (d): Bleomycin-intoxicated and CEE–treated group viewing weak Caspase-3 activity.

Fig. 7.

For a positive Caspase 3 immune reaction, intensity amounts are measured as mean ± SE * is considerably diversified in comparison to the control treatment, and # is significantly diversified with the bleomycin-treated rats (p ≤ 0.05).

4. Discussion

Many anticancer medications risk causing severe cardiotoxicity as a side effect that builds up over time and is dose-dependent. This risk exists for patients receiving treatment and healthcare professionals handling antiblastic medications. In reality, numerous investigations have demonstrated that certain cardiotoxic medicines, such as bleomycin, doxorubicin, epirubicin, cyclophosphamide, and 5-fluorouracil, were frequently found in the urine of exposed personnel.³⁰ Cardiotoxicity symptoms include arrhythmias, cardiomyopathy, and minor blood pressure changes.³¹Anticlastic medicines cause cardiotoxicity by producing free oxygen radicals, which cause cellular impairment, and stimulating immunogenic reactions due to antigen-presenting cells in the heart.³⁰.

LDH, CK-total, and CK-MB activities were significantly increased in adult male rats after receiving bleomycin 15 mg/kg twice a week, IP; these results are consistent with earlier studies that indicate bleomycin-induced oxidative stress might cause lipid peroxidation and release of these enzymes into the serum³². Bleomycin also caused cytotoxicity, apoptosis, and inflammation, as seen by the rise in LDH, CK-total, and CK-MB activities.².

The bleomycin treatment significantly increased cardiac NO and MDA levels relative to the untreated one and decreased cardiac GSH, GPx, SOD, and CAT. These outcomes are consistent with³³ and.4, 34 In our illustration, bleomycin led to an oxidative stress condition in the heart, as indicated by a decline in SOD activity and GSH level and an increase in NO activity. A key pathophysiological mechanism for heart injury is oxidative stress.³⁵ Internal antioxidant glutathione protects the heart from internal or external cardiac toxins.³⁶ The enzyme SOD catalyzes the formation of H₂O₂ from superoxide-free radical anions (O₂).³⁷.

Bleomycin could bind Fe (II) ions and create a complex that, when exposed to O₂, oxidizes to Fe (III), producing ROS such as superoxide, hydroxyl, and Fe (III) radicals. This bleomycin complex can bind nucleophilically with the DNA helix, breaking DNA strands, peroxiding membrane lipids, and ultimately damaging membranes.38, 39 In particular, the activity of the antioxidant system in the heart is weaker than in other tissues. Therefore, cardiomyocytes are vulnerable to attack by free radicals. Bleomycin has also been found to increase NO levels via an increase in inducible nitric oxide synthase (iNOS) expression⁴⁰. Increasing oxidative stress and overproduction of nitric oxide are implicated in the pathogenesis of cardiovascular diseases.⁴¹.

In the current study, oral treatment of ethanolic extract of costus for six weeks significantly raised the action of antioxidant defensives such (CAT, SOD, GSH, and GPx) in cardiac tissue but lowered MDA and NO accumulation, a lipid peroxidation indicator. These outcomes agree with those of.20, 42, 43 In this line,⁴⁴ The Costaceae plant species Costus pictus, a relative of S. costus, contained two chief polysaccharides (SLT-3 and SLT-4) that effectively decreased ROS production, controlled GPx activities, and hindered M.D.A. creation. Additionally, S. costus can contribute electrons to reactive radicals, turning them into more constant and unreactive classes, as demonstrated by.⁴⁵ According to research by,⁴⁶ the traditional natural antioxidant tocopherol is comparable in strength to the preventive activities of S. costus extract in protecting against toxicant-induced oxidative stress and the depletion of marker enzymes. The high levels of flavonoids, phenolic acids, steroids, and chlorogenic acid in this action may be linked to decreased membrane fluidity and deterioration.⁴⁷ Additionally, it has been demonstrated that prominent bioactive from S. costus, such as dehydrocostus lactone and costunolide, play significant roles as antioxidants by conjugating with particular proteins and mercapto (S.H.)-groups to influence several important biological functions in cells.⁴⁸ The plant's capacity to protect tissue macromolecules from the detrimental impacts of ROS-intermediated lipid peroxidation may account for the lower MDA concentrations in the Saussurea costus-treated group.⁴⁹.

High blood cholesterol levels, which are also considered one of the principal menace issues for myocardial infarction, speed up the development of atherosclerosis.⁵⁰ The Bleomycin injection resulted in hyperlipidemia and markedly elevated blood lipid levels. In experimental hypertriglyceridemia, triglyceride absorption from the systemic circulation is reduced due to decreased lipoprotein lipase action in the myocardium in bleomycin-injected rats. These findings concur with the investigation.³⁴ According to several authors⁵¹ The protective role of the S. costus roots may be explained by their high content of active ingredients with strong anti-inflammatory and antioxidant properties. This may also explain the observed improvement in lipid profiles, particularly the costus ethanolic extract.

In the existing investigation, intraperitoneal injection of bleomycin only resulted in a substantial increase in inflammatory protein levels (IL-6, IL-4, IL-10, and IL-1β) and a significant drop in CD4. These results support the findings.25, 44, 52 The interaction of bleomycin with iron and DNA produces ROS and initiates the inflammatory process.³⁶ Cardiovascular cells liberate inflammatory arbitrators and cytokines such as IL-4, IL-6, IL-1, TNF-α, and IL-10 in response to the ROS brought on by bleomycin. TNF-α synthesis impacted the mitochondria's ability to produce ROS and the inflammatory cells' creation of TNF-α and IL1β.⁵³.

Accumulating evidence has indicated that abnormalities in the production or function of cytokines, such as the pro-inflammatory cytokines TNF-α and IL-1beta, play a fundamental role in many inflammatory lesions.⁵⁴ Bleomycin has been reported to elevate ROS and trigger the production of TNF-α and IL-1β by creating the DNA/Fe2+/BLM complex.⁵⁵ The increased release of these cytokines causes an increase in TGF-β1 expression.⁵⁶.

The results show the extract's biological safety by lowering levels of cytokines and chemokines linked to inflammation, such as TNF-α, IL-6, IL-10, IL1β, and IL-4, in adult male albino rats after costus administration. In contrast to the standard treatment, these verdicts are consistent with those.⁴² Because it decreases the pro-inflammatory attributes such as TNF-α, IL-6, inducible NO synthase (NOS), and COX-2 (Cyclooxygenase) in numerous in vivo and in vitro trials, the latter plant has been identified as one of the plants with strong anti-inflammatory effects.⁵⁷ Helper CD4 T cells modify the situation and the action of other immune cells by secreting cytokines.³⁷.

The study of the cardiac myocytes histo-pathologically revealed a notable improvement. In contrast to rats treated with bleomycin, it kept the morphology of the heart muscle and decreased fibrosis and vacuolar degeneration. CEE's anti-inflammatory and antioxidant properties were said to be responsible for these results. The present findings were consistent with Ashry's investigation into the effectiveness of CEE in reducing cardio-hematotoxicity associated with Oxaloplatin therapy, in which he reported that CEE. had a protective role against Oxaloplatin in cardiac muscle tissue.⁵¹ Caspase3 expression is increased in experimental animals that experience heart failure and apoptosis, and people who have end-stage heart failure have activated caspase3 in their myocardium.⁴⁸ Caspase3 has been suggested as a marker for cardiotoxicity.⁵⁸.

5. Conclusions

According to the current study, post-treatment with S. costus extract may mitigate bleomycin-induced heart injury by reducing oxidative stress and inflammation that damage DNA and membranes (Fig. 8) These positive effects could result from these supplements' direct free radical scavenging properties. Therefore, the current study could recommend S. costus extract as supplemental medication to minimize the cardiotoxicity associated with the anticancer drugs.

Fig. 8.

Schematic diagram depicting the Cardioprotective, antioxidant, and anti-inflammatory efficacy of Coustus ethanolic extract against Bleomycin-Induced Cardiotoxicity in rats.

Ethics approval.

The Faculty of Science at Al-Azhar University in Assiut, Egypt's Animal Care and Use Committee provided all animals with humane treatment following its rules. The Faculty of Science, Al-Azhar University, Assiut, Egypt, which ethically approved the proposal with a number AZHAR 16/2023.

Consent to participate

All the authors agree to participate in this paper.

Consent for publication

All the authors agree with the publication of this paper.

CRediT authorship contribution statement

Barakat M. Alrashdi: Writing – review & editing, Resources, Data curation. Hussam Askar: Writing – review & editing, Supervision, Investigation, Formal analysis, Data curation, Conceptualization. Mousa O. Germoush: Formal analysis, Data curation. Maged Fouda: Writing – original draft, Formal analysis. Diaa Massoud: Writing – review & editing, Formal analysis, Data curation. Sarah Alzwain: Formal analysis, Data curation. Naser Abdelsater: Writing – review & editing, Formal analysis, Data curation. Laila M.S. Salim: Formal analysis, Data curation. Mohamed H.A. Gadelmawla: Writing – original draft, Formal analysis, Data curation. Mahmoud Ashry: Writing – review & editing, Supervision, Investigation, Formal analysis, Data curation, Conceptualization.

Declaration of competing interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Appendix A. . List of acronyms

Acronyms	Full Form
(iNOS)	inducible nitric oxide synthase
BLM	Bleomycin
Caspase 3	Cysteine-aspartic acid protease 3
CAT	Catalase
CD4	Cluster of Differentiation 4
CEE	costus ethanolic extract
CK- total	Creatine kinase-total
CK-MB	Creatine kinase-MB
COX-2	Cyclooxygenase
DNA	Deoxyribonucleic acid
Fe (II) ions	iron (II) or ferrous ion
Fe (III),	iron (III) or ferric
GPx	Glutathione Peroxidase
GSH	Reduced glutathione
H2O2	Hydrogen peroxide
HDL	density high-density lipoprotein
HPLC	High-performance liquid chromatography
IHC	Immunohistochemistry
IL-10	Interleukin-10
IL-1β	Interleukin-1beta
IL-4	Interleukin-4
IL-6	Interleukin-9
LDH	lactic acid dehydrogenase
LDL	Low-density lipoprotein
MDA	Malondialdehyde
NO	Nitric oxide
O2	Dioxygen
ROS	Reactive oxygen species
RSA	Radical Scavenging Activity
S. costus	Saussurea costus
S. verticillus	Streptomyces verticillus
SOD	Superoxide dismutase
TGF-β1	Transforming growth factor-beta 1
TNF-α	Tumor necrosis factor-alpha