Regulatory effect of Balanites aegyptiaca ethanol extract on oxidant/antioxidant status, inflammatory cytokines, and cell apoptosis gene expression in goat abomasum experimentally infected with Haemonchus Contortus

This experiment aimed to assess the regulatory effects of treatment with Balanites aegyptiaca fruit ethanol extract (BA-EE) on oxidant/antioxidant status, anti-inflammatory cytokines, and cell apoptosis gene expression in the abomasum of Haemonchus contortus–infected goats. Twenty goat kids were assigned randomly to four equal groups: (G1) infected-untreated, (G2) uninfected-BA-EE-treated, (G3) infected-albendazole-treated, (G4) infected-BA-EE-treated. Each goat in (G1), (G3), and (G4) was orally infected with 10,000 infective third-stage larvae. In the fifth week postinfection, single doses of albendazole (5 mg/kg.BW) and BA-EE (9 g/kg.BW) were given orally. In the ninth week postinfection, the animals were slaughtered to obtain abomasum specimens. The following oxidant/antioxidant markers were determined: malondialdehyde (MDA), glutathione (GSH), glutathione-S-transferase (GST), superoxide dismutase (SOD), catalase (CAT). The mRNA gene expression of cytokines (IL-3, IL-6, IL-10, TNF-α) and cell apoptosis markers (Bax, Bcl-2) were estimated. (G1) showed significantly reduced GSH content and GST and SOD activities but a markedly increased MDA level. (G3) and (G4) revealed a markedly lower MDA level with pronouncedly elevated GSH, SOD, and GST levels. The antioxidant properties of BA-EE were superior to those of albendazole. The mRNA gene expressions of IL-3, IL-6, IL-10, TNF-α, and Bax-2 were upregulated in (G1) but downregulated in (G3) and (G4). Bcl-2 and Bcl-2/Bax ratio expression followed a reverse course in the infected and both treated groups. We conclude that BA-EE treatment has a protective role in the abomasum of H. contortus–infected goats. This could be attributed to its antioxidant properties and ability to reduce pro-inflammatory cytokines and cell apoptosis.


Introduction
Haemonchus contortus (H. contortus), a highly pathogenic and voracious blood-feeding nematode, is a crucial gastrointestinal nematode (GIN) in small ruminants (Besier et al. 2016).The presence of this parasite causes economic losses because it reduces productivity, increases mortality rates, and requires expensive treatment (Arsenopoulos et al. 2021).H. contortus has a high propensity to develop resistance to anthelmintic drugs because of their overuse (Kotze and Prichard 2016;Bihaqi et al. 2020;Kaplan 2020).
During the GIN life cycle, excretory/secretory (ES) products are produced during the developmental processes to reach the adult stage (Besier et al. 2016).Many ES substances of H. contortus, including arginine kinase, galectins, and elongation factor 1 alpha proteins, have been found to play an essential role in modulating a wide range of biological functions, including cell activation, proliferation, adhesion, and apoptosis (Sun et al. 2007;Ehsan et al. 2017Ehsan et al. , 2020)).
The inflammatory response (inflammation) is one of the most vital defense mechanisms against numerous disease conditions and is associated with possible or actual tissue damage.The principal mechanisms involved in the inflammatory reaction are intimately linked to the production of free radicals and the induction of oxidative stress induced during the aggressive activity of GINs in the gastrointestinal tract, activating multiple immune mechanisms (Gadahi et al. 2016;Estrada-Reyes et al. 2017).Excessive levels of intracellular reactive oxygen species (ROS) frequently lead to the oxidative alteration of vital biomolecules, which can hasten cell apoptosis and exacerbate the disease condition (Lennicke and Cochemé 2021;Sies et al. 2022;Wang et al. 2023).The inflammatory processes activated by haemonchosis comprise large alterations in the production of cytokines, either in the peripheral blood or in the abomasum wall (Gossner et al. 2013;Estrada-Reyes et al. 2017).The host immune reactions against GINs involve an augmentation in cytokine production and the generation of antibodies specific to the parasite (McRae et al. 2015).These reactions are linked to decreased egg production, excretion by adult worms, and decreased parasitic burden as well as regulation of the damage caused by parasite burden (Machín et al. 2021).
The emerging anthelmintic resistance, drug toxicity, and customer demand for chemical-free animal products have raised the interest in natural alternative sources of anthelmintic drugs (Besier et al. 2016;Jaheed et al. 2019).Medicinal plants contain a wide spectrum of biological functions such as immunomodulatory, antioxidant, anti-inflammatory, and antimicrobial properties that have been attributed to polyphenols (Xu et al. 2017;Yahfoufi et al. 2018).Medicinal plants may promote local immune responses and improve the secreted cytokine profile, increasing 'resistance to haemonchosis in small ruminants (Hoste et al. 2016).Among the evidence-based herbal remedies is Balanites aegyptiaca (L.) Del. (B.aegyptiaca), often known as desert date in English and lalob, hidjihi, and heglig in Arabic.It is a member of the Zygophyllaceae family (Murthy et al. 2021).The fruits of B. aegyptiaca contain various kinds of compounds that possess a diverse range of pharmacological and biological activities, including antiparasitic, antioxidant, antibacterial, anti-inflammatory, and cytotoxic activities (Jaheed et al. 2019;Murthy et al. 2021;Sedky et al. 2022).Polyphenols (phenolic acids, flavonoids, and coumarins), alkaloids, steroids, saponins, and glycosides are just a few of the secondary metabolites that B. aegyptiaca tissues contain (Meda et al. 2011;Jaheed et al. 2019;Sedky et al. 2022).These fruits also contain minerals, major fatty acids, amino acids, and vitamins (Murthy et al. 2021).Therefore, B. aegyptiaca could be a source of natural antioxidant and anti-inflammatory compounds that may be extremely valuable as therapeutic agents for the prevention and treatment of various diseases (Speroni et al. 2005).In our previous study, we found B. aegyptiaca fruit extracts to have anthelmintic effects on some multiple-drug-resistant gastrointestinal parasites including H. contortus, both in vitro (Shalaby et al. 2020) and in vivo (for treatment) (Jaheed et al. 2019(Jaheed et al. , 2020)).B. aegyptiaca fruits are commonly used to purge intestinal parasites and have been found effective against H. contortus in goats (Jaheed et al. 2019), Fasciola gigantica in goats (Koko et al. 2000), Schistosoma mansoni in mice (Koko et al. 2005) and Trichinella spiralis in rats (Shalaby et al. 2010).
Based on our prior studies, B. aegyptiaca fruit ethanol extract (BA-EE) was abundant in phytochemicals, had a significant anthelmintic effect against H. contortus as evidenced by a decrease in fecal egg counts and worm burden of the parasite, and enhanced the health condition and metabolic profile of experimentally infected goats (Jaheed et al. 2019(Jaheed et al. , 2020)).Consequently, the purpose of this study was to investigate the regulatory influence of BA-EE treatment on oxidant/antioxidant status along with the anti-inflammatory cytokines and cell apoptotic and antiapoptotic gene expression profiles in the H. contortus-infected goats' abomasa.

B. Aegyptiaca fruit procurement and BA-EE preparation
B. aegyptiaca fruits were obtained from a local market in Aswan, Upper Egypt.Scientifically validated and voucher specimens were deposited in the Herbarium of Medicinal and Aromatic Plants Department, National Research Centre, Egypt.
B. aegyptiaca fruits were extracted using the approach provided by El-Newary et al. (2016).Briefly, the fresh fruits' mesocarp of B. aegyptiaca (1 kg) was exposed to triple maceration, soaked in 4 L of 70% ethyl alcohol, and maintained in tightly sealed jars at room temperature for three weeks, during which it was stirred several times daily with a sterile glass rod.This combination has been filtered.The residue was extracted 3-5 times in the same way until a clear, colorless supernatant extraction liquid was produced.The extracted liquid was filtered and concentrated with a rotary evaporator (Heidolph 2000, Germany) under decreased pressure at 50 °C until the solvent was totally evaporated.The extract was kept at 4 °C until further use.

Preparation of a dose BA-EE
BA-EE at a single dose of 9 g/kg.BW (body weight) was freshly suspended in 250 mL distilled water in a large bottle and drenched orally to the treated goats (Koko 2000;Albadawi 2010).

Albendazole
The anthelmintic efficacy of BA-EE was compared with that of albendazole (Evazole®, in the form of 2.5% oral suspension), a broad-spectrum anthelmintic which was used as a reference drug for treating one of the infected groups.Albendazole was purchased from the Veterinary Division of EVA Pharma, Cairo, Egypt.

Generation of the infective larval dose of H. Contortus
In order to develop infective third-stage larvae (L3) of H. contortus, an appropriate fecal culture was made using eggs from the collected worms (Soulsby 1982).A goat kid that tested negative for GIN infection was used as a donor, received 10,000 infective L3 in 10 ml of physiological saline solution orally, and served as a source of a monospecific L3 infection of the goats in the experimentally infected groups (Howlader et al. 1997;Jaheed et al. 2019).

Animals and experimental approach
Twenty male Egyptian Baladi goat kids (Capra hircus), aging six to nine months, weighing 18 to 21 kg, being free of internal parasites, and appearing healthy, were used.These animals were maintained in an intensive system of management in nematode-free settings for one month before the experiment.On the basis of zero nematode fecal egg counts, the kids were randomly divided into four equal groups of five each (G1-G4).Each animal in (G1), (G3), and (G4) received 10,000 infective L3 orally on the first day of the trial.Goats of (G1) (infected-untreated) were regarded as the control positive group.In the fifth week postinfection, goats of (G3) (infected-albendazole-treated) were treated orally with albendazole at a single dose of 5 mg/kg.BW (as a reference drug, EVA Pharma, Egypt) while goats of (G2) (uninfected-BA-EE-treated as control) and (G4) (infected-BA-EE-treated) were drenched orally with BA-EE at a single dosage of 9 g/kg.BW suspended in 250 mL distilled water.The trial lasted nine weeks.Each group was maintained in its own enclosure.

Abomasum tissue sampling
In accordance with the recommendations of the 'Ethical Committee for Medical Research (ECMR) of the National Research Centre (NRC), the animals were slaughtered at the end of the experiment.The abomasum specimens were quickly removed, cleaned with 0.9% ice-cold NaCl, and then kept at − 80 °C until used to measure antioxidant/oxidant markers, gene expression of inflammatory cytokines, and apoptotic markers.

Preparation of an abomasum homogenate
A homogenate of 10% (w/v) was produced by homogenizing one gram of abomasum tissue in an ice-cold 1.15% potassium chloride solution in a 50 mmol/L potassium phosphate buffer solution (pH 7.4).The homogenate was centrifuged at 4,000 ×g for five minutes at 4 °C.The supernatant was taken and stored at − 80 °C until used to determine antioxidant/oxidant markers.

Assessment antioxidant/oxidant markers in abomasum homogenate
Reduced glutathione (GSH) (Ellman 1959) and malondialdehyde (MDA) (Ohkawa et al. 1979) concentrations as well as the superoxide dismutase (SOD) (Marklund and Marklund 1974) activity were measured in the supernatant of the abomasum homogenate using a kit purchased from Bio-diagnostic Co., Egypt.Activities of glutathione-Stransferase (GST) (Habig et al. 1974) and catalase (CAT) (Aebi 1984) were chemically estimated.All analytical chemicals used were obtained from Sigma-Aldrich, USA.All parameters were measured using a spectrophotometer (T80 UV-Vis, PG Instruments Ltd., UK).

RNA extraction and quantitative real-time PCR
Total RNA was extracted from the abomasum tissue using TRIzol (Invitrogen, Carlsbad, CA, USA) in accordance with the manufacturer's instructions.The yield and quality of the RNA were analyzed using a NanoDrop™ 1000 spectrophotometer and gel electrophoresis (Thermo Fisher Scientific, USA).RNA (1 µg) was treated with an RNase-free DNase kit (Promega) to remove any genomic DNA contamination, and cDNA was synthesized using a reverse kit (RT-PreMix Kit for cDNA synthesis, iNtRON Biotechnology, Seongnam-Si, Korea).The reaction was carried out in a thermal cycler under specific cycling conditions: 42 °C for 60 min and 90 °C for 5 min.
The data are represented using the 2 −ΔΔCt method as the fold change in target gene expression normalized to the housekeeping genes β-actin and GAPDH to normalize input RNA quality, RNA quantity, and reverse transcription efficiency.

Statistical analysis
All data were statistically analyzed and presented as means ± standard error (SE).Differences between data of antioxidant/oxidant parameters in different groups were tested for significance by the one-way analysis of variance References IL: Interleukin.TNF-α: Tumor necrosis factor alpha.Bax: B-cell lymphoma-2 protein-associated X protein Bcl-2: B-cell lymphoma-2 protein.GAPDH: Glyceraldehyde-3phosphate dehydrogenase formation during the reaction.Each sample's quantitative gene expression was estimated, and the Ct means were calculated.IL-3 is a pleiotropic hematopoietic cytokine.Its quantitative mRNA gene expression was significantly (P ≤ 0.0001) upregulated in the infected-untreated goats compared with the uninfected-BA-EE-treated ones.Treatment with albendazole or BA-EE markedly downregulated the mRNA gene expression levels, with a significant difference between their values (− 3.1-and − 2.3-fold change; P = 0.005, P ≤ 0.000; respectively; Fig. 1a).
The cytokine IL-6 has a pleotropic role in controlling inflammation and the immune system.The relative mRNA gene expression of IL-6 was markedly (P = 0.0003) upregulated in the infected-untreated goats compared with the uninfected-BA-EE-treated ones.Furthermore, treatment with either albendazole or BA-EE downregulated the mRNA gene expression levels, with no significant difference between their values (− 2.5-and − 2.1-fold change; P = 0.004, P = 0.0002) in the infected-albendazole-treated and infected-BA-EE-treated groups, respectively, in comparison with the infected-untreated group (Fig. 1b).
The cytokine IL-10, which has strong anti-inflammatory properties, is essential for limiting the response of the host immune system' to pathogens.The relative mRNA gene expression of IL-10 was significantly (P = 0.001) upregulated in the infected-untreated goats compared with the uninfected-BA-EE-treated ones, whereas treatment with either BA-EE or albendazole reduced fold expression with no significant difference (P = 0.788; Fig. 1c).
The pro-inflammatory TNF-α cytokine is released by macrophages and regulates several cellular processes, such as cell division, proliferation, apoptosis, lipid metabolism, and coagulation.Gene expression results indicated a significant (P ≤ 0.0001) fold increase in the infected-untreated group compared with the uninfected-BA-EE-treated one.Meanwhile, treatment with either BA-EE or albendazole decreased fold expression, with no significant difference between their values (P = 0.785; Fig. 1d).
(ANOVA) followed by Duncan's multiple range test using the SPSS version 17 (SPSS Inc., Chicago, IL, USA) computer program.Statistical analyses of data of gene expressions of inflammatory cytokines and apoptosis were conducted with Prism v8.3.1 (GraphPad, La Jolla, CA, USA).Differences between groups were evaluated by oneway ANOVA, and the significant level was set as P ≤ 0.05.

Antioxidant/oxidant status
The data presented in Table 2 illustrate the effect of BA-EE and albendazole (as a standard drug) on the levels of oxidant and antioxidant parameters in the abomasum homogenate of goats experimentally infected with H. contortus.
Goats infected with H. contortus and untreated revealed that GSH content and GST and SOD activities significantly (P < 0.001) decreased while the MDA level markedly (P < 0.001) increased in the abomasum homogenate compared with the other groups.After BA-EE and albendazole treatment, the GSH content and SOD and GST activities markedly increased while the level of MDA significantly decreased in comparison with the infected-untreated goats.After treatment with BA-EE, CAT markedly (P < 0.05) elevated in comparison with the other groups.There were significant differences between all parameters' values after treatment with BA-EE and albendazole, which revealed that BA-EE had greater antioxidant effect than albendazole (Table 2).

Cytokine gene expressions
A quantitative PCR was carried out to estimate the changes in mRNA gene expressions of cytokines (IL-3, IL-6, IL-10, and TNF-α) in the abomasum tissue of the infected goats compared with the other groups, and β-actin and GAPDH were used as housekeeping genes.All of the tested genes showed a single peak in the PCR melting curve, indicating the specificity of amplification and the lack of primer-dimer the expression levels of these genes and the ratio of Bcl-2 to Bax.The mRNA content of Bax in the abomasum tissue of the infected-BA-EE-treated goats dropped by 47% compared with the infected-untreated group (P = 0.0001; Fig. 2a).Moreover, we found that Bcl-2 gene expression increased by 21% in the abomasum tissue of infected-BA-EE-treated goats compared with the infected-untreated ones (P = 0.0001; Fig. 2b).Additionally, the ratio of Bcl-2 to Bax in the abomasa of the infected-untreated goats (21%) was considerably lower (P = 0.0001) than that in the uninfected-BA-EE-treated ones (Fig. 2c).

Discussion
Because of the highly anthelmintic efficacy of BF-EE in our previous study (Jaheed et al. 2019), a maximal reduction in eggs per gram of feces (EPG) (88.10%) and worm burden (94.66%) was recorded in the fourth week posttreatment in contrast to the efficacy of albendazole (98.29% and 96.95%, respectively).Also, BF-EE improved the health condition and metabolic profile of infected goats.In our previous work (Jaheed et al. 2019), the phytochemical components screened included saponins, alkaloids, flavonoids, phenolics, and terpenoids.In addition, the crude BA-EE exhibited

Apoptotic gene expression
The quantitative gene expression of proapoptotic Bax was significantly (P ≤ 0.0001) upregulated in the infected-untreated goats compared with the uninfected-BA-EE-treated ones.Furthermore, treatment with either albendazole or BA-EE markedly downregulated the expression levels (− 1.7-and − 1.5-fold change; P = 0.0007, P = 0.0003, respectively).There was no statistically significant difference between the infected-BA-EE-treated and uninfected-BA-EE-treated groups (P = 0.167).Also, there was no significant change (P = 0.752) in Bax expression in the infected-albendazoletreated or infected-BA-EE-treated group (Fig. 2a).
The expression of the antiapoptotic gene Bcl-2 in the infected-untreated group showed a downregulation that was nonsignificant compared with that in the uninfected-BA-EE-treated group.Furthermore, treatment with either albendazole or BA-EE upregulated the expression levels (2.3-and 3.1-fold change; P = 0.0003, P = 0.0001, respectively; Fig. 2b).

Bcl-2/Bax ratio
In terms of the expressions of the Bax and Bcl-2 genes, it was found that the capacity of antiapoptotic Bcl-2 to suppress the activity of proapoptotic Bax was impacted by both GSTs are a protein family that catalyzes the conjugation of GSH to various electrophiles via the sulfur atom of cysteine (Salinas and Wong 1999).Similarly, we found that H. contortus infection caused a decreased GST activity in the abomasum tissue, which is somewhat consistent with the observation made by Bártíková et al. (2010), who showed that intestinal GST activity was not significantly decreased after sheep suffered from haemonchosis.
Also, a significant decrease in hepatic GST activity was observed in male mouflons (Skálová et al. 2007) and Mouflon ewes infected with Dicrocoelium dendriticum (Křížová et al. 2008) and in the ileum of rats infected with Cryptosporidium (Abd El-Aziz et al. 2014).In contrast, an increase in GST activity was observed in goats spontaneously infected with haemonchosis (Rashid and Irshadullah 2019).The reason for the different influence of H. contortus infection on the GST activity may be the worm burden as reported by Rashid and Irshadullah (2019), who studied the effect of infection in naturally infected goats.
On the other hand, the infected abomasum tissue showed a significant decline in SOD activity, which is consistent with the findings of Rashid and Irshadullah (2014), Ahmed et al. (2019), andAlam et al. (2020).Similarly, the lower SOD activity has been reported in ovine naturally infected with Fasciola hepatica (Kolodziejczyk et al. a strong in vitro total antioxidant activity (2128.86mg/100 g ascorbic acid equivalent).
MDA, as a lipid peroxidation by-product, is regarded as a good indicator for cell damage, which confirmed the increased release of free radicals in the H. contortus-infected animals (Ayala et al. 2014).MDA was markedly elevated in the abomasum tissue after H. contortus infection (Jaheed et al. 2020;Mravčáková et al. 2021), which is consistent with the present result.Reduced GSH (γ-L-glutamyl-Lcysteinyl-glycine) is a tripeptide thiol that helps in eliminating the cytotoxic effect of ROS and protecting cells from it.H. contortus infection was accompanied by a reduction in the levels of GSH in the abomasum tissue, which were comparable to those reported by Rashid and Irshadullah (2019) and Ahmed et al. (2019).According to the hematophagous activity of both adult and fourth larval stage of H. contortus, the decreased GSH level could be due to a decrease in the precursor of the amino acid levels (cysteine and cystine) responsible for GSH synthesis (Sido et al. 1998).Moreover, the antioxidant function of GSH is accomplished largely by using the GSH as a substrate for GPx, which detoxifies peroxides to water, while two GSH molecules are oxidized to form oxidized GSH (GSSG).So, the reduced level of GSH may also be explained by the increased oxidation of GSH to GSSG, catalyzed by free radicals (Morris et al. 2013).Similar findings have been reported in mice infected with competing with the apoptotic factor Bcl-2. Apoptosis is regulated by the balance between Bax and Bcl-2 expression (You et al. 2019;Zheng and Kanneganti 2020).
The current results of apoptotic gene expression patterns revealed higher levels of Bax, decreased levels of Bcl-2, and changes in the ratio of Bcl-2 to Bax in the infected abomasum tissue.Accumulating evidence suggests that the infected abomasum tissue is mediated by apoptotic signaling pathways, which are also connected to ROS-induced oxidative damage (Sies et al. 2022).This is supported by our findings that H. contortus causes an increase in the MDA levels and depletion in the antioxidant state of the infected animals, which is represented by lowered GSH, GST, and SOD levels.Lipid peroxidation, a process that directly degrades phospholipids, is utilized as a cell-death signal to induce programmed cell death (Lennicke and Cochemé 2021).
Furthermore, the recombinant galectins of the H. contortus parasite (rHco-gal-m/f proteins) were shown to trigger apoptosis in goat peripheral blood lymphocytes (PBLCs) (Sun et al. 2007).Ehsan et al. (2017;2020) suggested that two key ES proteins of H. contortus, arginine kinase and elongation factor 1 alpha (rHcEF-1α), inhibited the proliferation and enhanced the apoptosis of goat peripheral blood mononuclear cells (PBMCs) in vitro.According to La Sala et al. (2001), arginine kinase controls the transformation of adenosine triphosphate (ATP) molecules into arginine, which is necessary for metabolic and cellular processes such as cytokine production, cellular response, plasma membrane alteration, and apoptosis.
In the present study, the infected goats treated with BA-EE showed an increase in the antioxidant levels (GSH, SOD, GST, and CAT) accompanied by a decrease in the MDA level.This could be because crude BA-EE exhibited a strong antioxidant activity in vitro (Jaheed et al. 2019).Furthermore, BA-EE reduced the production of proinflammatory cytokines, including TNF-α, IL-3, and IL-6; strengthened its anti-inflammatory role (Gaur et al. 2008;Elkareem et al. 2021); and regulated apoptosis through the balance between Bax and Bcl-2, the two markers of apoptosis gene expression.This could be due to either the antioxidant activity (Jaheed et al. 2019;Sedky et al. 2022) or the anthelminthic efficacy of BA-EE which caused the reduction of worm burden (Jaheed et al. 2019;Shalaby et al. 2020).
As was previously mentioned, the anti-inflammatory properties of BA-EE may be responsible for its inhibition of the release of the inflammatory mediators IL-6 and TNF-α with enhancement of production of the anti-inflammatory IL-10, as well as antioxidant properties of BA-EE (Gaur et al. 2008;Elkareem et al. 2021).
B. aegyptiaca fruit extracts have been shown to contain phytochemical components such as phenolic compounds 2005), Theilaria sp.(Nazifi et al. 2011) and Babesia ovis (Esmaeilnejad et al. 2012).The inhibition of SOD activity in the infected abomasum tissue could be attributed to the overproduction of the superoxide anion which causes inactivation and depletion of SOD storage (Rashid and Irshadullah 2014).Generally, these findings indicated an enhanced oxidative stress with a decline in antioxidant defense in the H. contortus-infected goats.
In the present study, haemonchosis elevated IL-3, IL-6, IL-10, and TNF-α gene expressions in abomasum tissue mRNA, which was attributed to the activation of the inflammatory cascade triggered by infection.H. contortus infection increases pro-inflammatory cytokines because of the elevated intracellular ROS in the abomasum, which in turn stimulates the transcription factor nuclear factor kappa B (NF-κB) (Bąska and Norbury 2022).NF-κB activation triggers inflammatory signaling cascades and an increase in TNF-α expression in the abomasum (Toscano et al. 2019).TNF-α stimulates the production of inflammatory cytokines, particularly IL-3, IL-6, and IL-10 (Jang et al. 2021;Kany et al. 2019).These results agree with Terefe et al. (2007) which identified increased levels of IL-3 gene expressions in H. contortus-infected lambs.Meanwhile, Buendía-Jiménez et al. (2015) revealed increased IL-6 gene expression in sheep infected with H. contortus.Moreover, the same results were noticed in the tropical sheep breed Pelibuey infected with H. contortus (Estrada-Reyes et al. 2015, 2017).
Although apoptosis is a form of programmed cell death that is essential for proper tissue development, a variety of diseases including helminths are able to manipulate host apoptosis pathways to their benefit (Donskow-Schmelter and Doligalska 2005).Apoptotic proteins Bcl-2 and Bax govern cell death.Bcl-2 pore-stabilizing proteins maintain mitochondrial membrane barrier function, inhibiting apoptosis and prolonging cell survival.Bax is primarily found in cytoplasm where it functions as a pore-destabilizing protein the parasite.It also induced degenerative alterations in the parasite tegument and the cells damaging its metabolism, resulting in immobilization and death of the cyst (Jung-Cook 2012).
In conclusion, as a promising supplement, BA-EE has an anthelmintic effect exhibited by its protective role in the abomasum tissue of goats infected with H. contortus.It exhibited modulatory effects on oxidant/antioxidant profile and inflammatory cytokines, as well as up-and downregulatory effects on the mRNA gene expression levels of antiapoptotic and proapoptotic genes, respectively.This could be explained via the antioxidant and anti-inflammatory activities of its bioactive ingredients.
Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made.The images or other third party material in this (gentisic, p-coumaric, caffeic, ferulic, and sinapic acids), quinones, flavonoids, flavones, tannins, coumarins, terpenoids, essential oils, and alkaloids (Jaheed et al. 2019;Murthy et al. 2021;Sedky et al. 2022).Condensed tannins are the most important constituent of B. aegyptiaca and have an anthelminthic activity against H. contortus in vitro (Assefa et al. 2018).It has been shown that both ethanolic and aqueous extracts of B. aegyptiaca fruits exhibit significant antioxidant power in vitro (Jaheed et al. 2019;Sedky et al. 2022).The antioxidant and anti-inflammatory properties of B. aegyptiaca have been linked to its phenolic and flavonoid content (Yahfoufi et al. 2018;Murthy et al. 2021).Natural compounds, with their ability to modulate pro-inflammatory gene expression and antioxidant properties such as ROS scavenging, play a crucial role in regulating inflammatory signaling (Malireddy et al. 2012;Yahfoufi et al. 2018).These findings could be attributed to the bioactive BA-EE compounds, which induce different parasiticidal mechanisms.Previous publications have shown that BA-EE has promising anthelmintic efficacy (Jaheed et al. 2019;Shalaby et al. 2016Shalaby et al. , 2020;;Hassan et al. 2021).
Studies conducted in vivo and in vitro have shown that polyphenols have an impact on macrophages by suppressing a number of essential inflammatory response regulators, such as TNF-α, IL-1, and IL-6 (González et al. 2011).Additionally, flavonoids have been demonstrated to reduce the levels of a variety of pro-inflammatory cytokines, particularly IL-1, IL-6, IL-8, and TNF-α (Comalada et al. 2006).
Similarly, several polyphenol analogs, such as the curcumin analog EF31, have been demonstrated to decrease the secretion and expression of IL-1, IL-6, and TNF-α in mice (Olivera et al. 2012).Likewise, TNF-α and IL-6 production is reduced in THP1 macrophages by willow bark, meadowsweet, and extracts of chamomile and isolated polyphenols such as quercetin found in these extracts (Drummond et al. 2013).Also, the phytochemical analysis of BA-EE revealed the presence of α-linolenic acid as one of the major constituents.α-Linolenic acid is an omega-3 fatty acid known to possess an anti-inflammatory activity (Otuechere and Farombi, 2020), which may explain the reported inhibition of the pro-inflammatory mediators.
The treatment with albendazole enhanced the host immune response by changing the cellular infiltration and thus altering the cellular immunity (Ricken et al. 2017).Albendazole treatment in goats reduced H. contortus infection (87.9%) more than levamisole (80.2%) and parbendazole (83.9%) did (Charles et al. 1989).Administration of albendazole influenced the balance of immune response and promoted the secretion of pro-inflammatory factors, which is beneficial to parasite clearance (Wu et al. 2021).Albendazole treatment supported the activation of the host immune system by reducing the immunosuppressive functions of article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material.If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder.To view a copy of this licence, visit http://creativecommons. org/licenses/by/4.0/.