{"id":61073,"date":"2024-09-30T10:40:25","date_gmt":"2024-09-30T10:40:25","guid":{"rendered":"https:\/\/biomedpharmajournal.org\/?p=61073"},"modified":"2024-10-09T18:36:30","modified_gmt":"2024-10-09T18:36:30","slug":"the-effect-of-an-aquatic-extract-of-eucalyptus-globulus-leaves-on-reducing-the-inflammation-parameters-caused-by-carrageenan-in-male-wistar-rats","status":"publish","type":"post","link":"https:\/\/biomedpharmajournal.org\/staging\/vol17no3\/the-effect-of-an-aquatic-extract-of-eucalyptus-globulus-leaves-on-reducing-the-inflammation-parameters-caused-by-carrageenan-in-male-wistar-rats\/","title":{"rendered":"The Effect of an Aquatic Extract of Eucalyptus globulus Leaves on Reducing the Inflammation Parameters Caused by Carrageenan in Male Wistar Rats"},"content":{"rendered":"\n

Introduction<\/strong> <\/p>\n\n\n\n

In this study,\nwe investigated the anti-inflammatory properties of an aquatic extract of Eucalyptus\nglobulus <\/em>(ECP) by inducing inflammatory indices using carrageenan. Herbal\nproducts are essential sources of herbal extracts and other pharmacological\nagents. Since ancient times, people have used the leaves of various Eucalyptus<\/em>\nspecies as traditional medicines to treat fever, colds, and other ailments1<\/sup>.<\/p>\n\n\n\n

ECP leaves are\nutilized locally to treat burns, dermatitis, scabies, tonsillitis, and\ndysentery; they are also taken orally to treat articular pain, tonsillitis, dysentery,\ninfluenza, and cystitis. Research has shown that using ECP leaves orally can\nreduce cold symptoms during flu-episodes2,3<\/sup>. Eucalyptus globulus <\/em>possesses remarkable pharmacological\nactivities, including antioxidant and anti-inflammatory effects4<\/sup>. The naturally oils in Eucalyptus\nglobulus<\/em> include 1-eucalyptol, which has a variety of pharmacological\nactions, including insecticidal, gastrointestinal, dermatological,\nanti-inflammatory, analgesic, antidiabetic, antioxidant, anticancer, and\nantibacterial properties5\u20137<\/sup>.<\/p>\n\n\n\n

Concisely, Eucalyptus<\/em>\nleaves have the ability to protect the liver from damage and reduce oxidative\nstress due to reactive oxygen species (ROS) produced during metabolism8<\/sup>, It has previously been shown to be a source of bioactive\nchemicals, namely phenolic compounds like phenolic acids, flavonoids, or\nhydrolyzable tannins. E. globulus<\/em> leaves have long been used to treat\nrespiratory issues9,10<\/sup>.<\/p>\n\n\n\n

Eucalyptus<\/em> leaves have antioxidant properties, as evidenced by their ability\nto scavenge free radicals, activate antioxidant enzymes, and suppress\ninflammation by inhibiting lipoxygenase and lowering nitric oxide (NO) levels10<\/sup>. It was demonstrated that their ethanolic extracts reduced the\nlevels of pro-inflammatory mediators and nitric oxide (NO)11<\/sup>, and that they had the ability to scavenge free radicals12<\/sup>.<\/p>\n\n\n\n

A basic defense mechanism of the host in reaction to damaging\nstimuli such as, illness, and\/or irritation is inflammation13<\/sup>. The symptoms of acute inflammation manifest suddenly and include\nedema, leukocyte emigration, enhanced vascular permeability, and vasodilation14<\/sup>. A series of cytokines governs the inflammatory process to a\nsignificant extent and is essential for cell-to-cell contacts and communications15<\/sup>.<\/p>\n\n\n\n

Carrageenan\n(E407) is a polysaccharide extracted from red algae (Chondrus crispus) that\ninduces inflammation induced inflammatory in the rat16,17, Acute\ninflammation leads to the development of biphasic inflammatory responses in the\ntissue of the paw18. The main\nobjective of the current research is to ascertain the therapeutic properties of\nEucalyptus<\/em> leaves and their role in mitigating the inflammatory activity\nof carrageenan.<\/strong><\/p>\n\n\n\n

Research\ndesign and methods<\/strong><\/strong><\/p>\n\n\n\n

The current study aims to identify the therapeutic qualities of eucalyptus leaves and how they contribute to reducing carrageenan’s inflammatory activity. We obtained 24 male Albino rats, aged 12 to 14 weeks, from the Animal House, College of Pharmacy \/ University of Karbala. The Institutional Animal Ethics Committee, University of Kerbala, Iraq, assisted in reviewing and approving all animal experimentation protocols, with the approval of experimental research. Animal maintenance therapies, as instructed by animal care experts, adhere to the International Animal Care and Use Guidelines19<\/sup>. We randomly divided these rats into four groups, each containing six rats, kept them in a separate cage, and gave them two weeks to acclimate to the water and food supplies, following the American Institute of Nutrition’s semi-purified diet (AIN).<\/p>\n\n\n\n

Group 1: control, which receives an oral dose of 2 ml of a normal saline physiological solution for 30 days.<\/p>\n\n\n\n

Group 2: is the positive control\u060c receiving 0.6 g\/kg of Eucalyptus globulus<\/em> extract (ECP) orally at a dose of 2 ml for 30 days. The (LD\u2085\u2080) of eucalyptus <\/em>oil is 2770-4500 mg\/kg (rat)20<\/sup>.<\/p>\n\n\n\n

Group 3: For 30 days, the Carrageenan group will receive an intraperitoneal injection of 100 \u03bcl of the carrageenan solution (CRG) dissolved in 1 milliliter of distilled water.<\/p>\n\n\n\n

Group 4: CRG and ECP Group (C&E): For fifteen days, this group receives an intraperitoneal injection of 100 \u03bcl of the carrageenan solution dissolved in 1 milliliter, followed by an oral dose of ECP after two hours, to follow up on the inflammation status in the C&E group, in which inflammation was decreasing compared to the CRG group, in which inflammation was ascending (inflammation is directly proportional to the time of carrageenan injection) in the same period, which 30 days. We assumed that a period of 15 days is sufficient to stimulate an inflammatory state using intraperitoneal carrageenan21<\/sup>, followed by an equal period of treatment using ECP to try to restore inflammatory factors to their normal levels as in the control group.<\/p>\n\n\n\n

Following a\n12-hour fast, we sacrificed the rats after 30 days and extracted seven ml of\nblood directly from the heart for a complete blood count (CBC). We recorded the\nresults and separated the blood serum using an Inflammation Panel 1 Rat Kit to\nmeasure the percentage of oxidants, antioxidants, and inflammation. We assessed\nand contrasted the proportion of inflammation in the treated group with that of\nthe control group.<\/p>\n\n\n\n

Plant Materials\nCollection and Extraction Procedure<\/strong><\/p>\n\n\n\n

We collected\nthe Eucalyptus globulus<\/em> leaves from our garden in Karbala, Iraq, in\nFebruary 2024. The National Herbarium. We cut and divided the plant leaves into\nsmall pieces, dried them in the air for a few days, and ground them to powder\nusing a motorized grinder (beerfingo, SR01, Algeria). We used a weighing\nmachine (Suyue, China) to powder the plant materials into 250 g, which we then\nsubmerged in 750 ml of distillation water in airtight containers for three\ndays. We then filtered the powder through a fresh cotton bed and No. 2 filter\npaper, respectively. We stored the crude extracts at 25 \u00b0C in well-closed\ncontainers for future use. <\/p>\n\n\n\n

Devices and Substances Used in the Study<\/strong><\/p>\n\n\n\n

To prepare\nCarrageenan, type IV (Sigma, USA), dissolve 0.01 g in 1 ml of distilled water.\nWe measured the white blood cell (WBC) leucocyte, lymphocyte, monocyte, and\nneutrophil count by used BC-3000 hematology machines from Mindray (India). We\nused Rat CRP detection kits from Chondrex (USA) to measure the C-reactive protein\n(CRP) and a Rat TNF enzyme-linked immunosorbent assay kit (ELISA) for the TNF\nassay.<\/p>\n\n\n\n

Data Analysis<\/strong><\/p>\n\n\n\n

We display each result as the mean and standard deviation. ANOVA\nanalysis was utilized to determine the experimental data’s statistical\nsignificance, with a significance threshold of P <.05We used the Excel 2013\nprogram as a numerical instrument to carry out this statistical study.<\/strong><\/p>\n\n\n\n

Results<\/strong><\/p>\n\n\n\n

Tables 1 and 2 displays the noteworthy variations in blood\nparameter values between groups. Carrageenan has been altering the levels of\nWBC, lymphocytes, monocytes, neutrophils, CRP, and TNF. The levels of\nmonocytes, neutrophils, CRP, and TNF were all the same between the ECP and the\ncontrol groups (P = 0.05), but there was a remarkable rise (P< 0.05) between\nthe CRG and the C&E group in contrast to the group of control. Carrageenan\nhas caused significant Inflammation in the CRG group. The table also clearly\nshowed that Carrageenan impacted WBC, lymphocytes, monocytes, neutrophils, CRP,\nand TNF. The CRG group showed a remarkable rise in contrast to the control and\nECP groups. This result aligns with numerous previous studies22,23<\/sup>. This is due to the inflammation induced by carrageenan in the\nexperiment animals24,25<\/sup>, based on the findings of previous studies22,23,25<\/sup>. Additionally, Table 1 demonstrated that there was a considerable\nrise in the ECP group’s WBC count as a result of ECP (P< 0.05) in contrast\nto the control group and a significant decrease (P< 0.05) compared to the\nCRG and C&E groups. In other words, the ECP reduced the count of WBC in the\nC&E group in contrast to the CRG group and caused a slight increase in the\nECP group compared to the control group. The effect of ECP on lymphocyte count\nincreased in the ECP group as shown in Table 1, with a significant increase\n(P< 0.05) in contrast to the control group and a significant decrease (P<\n0.05) compared to the CRG and C&E groups. In addition, there are no\nsignificant differences. (P = 0.05) There is a significant difference (P =\n0.05) in monocyte counts between the ECP and control groups. Table 2 showed a significant increase\n(P< 0.05) in the level of CRP in the serum of rats belonging to the CRG\ngroup compared to the all-experimental group. There are no significant\ndifferences (P = 0.05) between the ECP and control groups. These results\nare consistent with Study26<\/sup>. Finally, Table 2 shows a descending order of TNF levels in rats’\nserum among the experimental groups. The highest level (243.50 \u00b1 18.59 Pg\/mL)\nwas recorded by Up, with highly significant differences (P< 0.05) compared\nto all other experimental groups, including the control group. It was followed\nby the treatment group that was given CRG for fifteen days, followed by the\ntreatment period with ECP for fifteen days (186.00 \u00b1 3.85 Pg\/mL), then the ECP\nand the control groups (133.50 \u00b1 3.51 Pg\/mL) and (127.83 \u00b1 8.33 Pg\/mL), respectively,\nbetween which there were no notable variations (P = 0.05).<\/p>\n\n\n\n

Discussion<\/strong><\/p>\n\n\n\n

The Effect of Carrageenan on the Blood Parameters Levels<\/strong><\/p>\n\n\n\n

According to\nexperimental data in Tables 1 and 2, CRG changes the microbiota, stimulates the\nrelease of pro-inflammatory cytokines, activates innate immunity pathways, and\ninitiates or intensifies the inflammatory response27<\/sup>.<\/p>\n\n\n\n

In general, WBC count increases in many inflammation conditions28<\/sup>, such COVID-1929<\/sup>, Parkinson’s disease30<\/sup>, rheumatism, lymphoid inflammation31<\/sup>, liver diseases32<\/sup>, and  urinary tract inflammation33<\/sup>, induced colitis34<\/sup>. In this study, increases of peripheral Leucocytes, that suggest because of carrageenan24<\/sup>, by stimulated macrophages and the release of IL-1\u03b2 that have a major part in the inflammatory reaction35<\/sup>. CRG can elicit an acute inflammatory response with polymorphonuclear neutrophils infiltration and exudate increasing in cells36<\/sup>.<\/p>\n\n\n\n

Carrageenan\nstimulated the production of cytokines\/chemokines by neutrophils or macrophages37<\/sup>, resulting tissue damage38<\/sup>, and this leads to estimate the extent of damage caused by\ncarrageenan39<\/sup>. Increase level of C-reactive protein (CRP)\nin the CRG group40<\/sup>.<\/p>\n\n\n\n

Moreover, CRG\nreported to affect the oxidative burst activity in neutrophils. The oxidative\nburst is a crucial process by which neutrophils eliminate ingested pathogens by\nthe formation of reactive oxygen species (ROS)41<\/sup>.<\/p>\n\n\n\n

It seems that\nthe mechanism responsible for this impact is the activation of certain\nsignaling pathways linked to inflammation. CRG attaches to immune cells’ toll-like\nreceptors (TLRs)42<\/sup>, setting off a\nseries of events that lead to increased neutrophil synthesis and recruitment.\nThis mechanism is necessary to create a successful immune response, but it can\nalso result in excessive inflammation if it is not control appropriately43<\/sup>. That demonstrate of induced inflammation44<\/sup>, as the best-known inflammatory marker in an oncological\nsituation, CRP, represents an inflammatory agent45<\/sup>.<\/p>\n\n\n\n

In addition to the WBC, lymphocyte, monocyte, neutrophils, and CRP,\nthe significant increase in the levels of TNF as inflammatory mediator in the\nCRG group it is consider another evidence of inflammation in this group34,46<\/sup>.  One of the most\neffective medications for the management of autoimmune and chronic inflammatory\ndiseases is a biologic that neutralizes TNF, a key cytokine in inflammatory\nresponses47<\/sup><\/p>\n\n\n\n

Inflammatory reactions can be triggered directly through the\nproduction of inflammatory genes or indirectly through the induction of cell\ndeath47<\/sup>. <\/p>\n\n\n\n

Consequently, it shown that in a range of\nanimal models of TNF-induced inflammatory diseases, genetically guided cell\ndeath may be able to reverse the inflammatory phenotype48<\/sup>. Cells do not\nalways react to TNF by dying. In order to shield the organism from possible\nharm, protective brakes, also known as cell death checkpoints, often actively\ninhibit TNF cytotoxicity, Although TNF can cause cell death, this reaction only\nhappens when one of the checkpoints for cell death deactivated49<\/sup>.<\/p>\n\n\n\n

The effect of ECP on WBC count, Lymphocytes, Monocytes and Neutrophils<\/strong><\/p>\n\n\n\n

In this study we investigation to determine the ant-inflammatory of\naqueous extract of Eucalyptus globulus<\/em> (ECP) by the inflammatory\nindices. The data of this study in the Table 1, it is attributed to the\npresence of active substances in the ECP that have a protective role against\ninflammatory factors caused by CRG50,51<\/sup>. Most cases of inflammation are accompanied by a significant\nincrease in the of WBC count52<\/sup>, This is because these cells have primary functions in eliminating\nforeign bodies such as bacteria53<\/sup>, cancer cells 54<\/sup>, and cell debris55<\/sup>. ECP has antibacterial56<\/sup>, anti-inflammatory properties57<\/sup>, therefore played a major role in reducing inflammation caused by\nthe effect of CRG in the C&E group.<\/p>\n\n\n\n

Lymphocytes and monocytes are types of WBC\nwith Polymorphonuclear, Lymphocytes play a major role in correlating at a cellular\nlevel with the accumulation in lymphoid tissues58<\/sup>. As an illustration in Table 1, the\nevidence for clonal expansion in innate lymphocytes\u2014which has largely been\nobserved in cytotoxic natural killer (NK) cells responding to CMV infection is\ngathered, in response to this viral infection, NK cells in particular go\nthrough clonal growth in a manner specific to each antigen on their route to developing\nmemory characteristics59<\/sup>.<\/p>\n\n\n\n

Aside from the fact that macrophages are the most prevalent immune cell\ntype in adipose tissue, other immune cells, such as B cells, are also present\nand play significant roles in regulating adipose tissue inflammation. These\nfactors explain the immune system’s involvement in obesity-induced adipose\ntissue inflammation and the ensuing metabolic dysfunction60<\/sup>.<\/p>\n\n\n\n

Since it proven\nin this study that CRG caused inflammation in experimental animals in the CRG\nand C&E groups, which was evident through the increase in the inflammatory\nindicators mentioned above, including WBC, monocytes and neutrophils as clear\nin the data of the Table 1, ECP make to maintain the integrity of the cells\nexposed to inflammation and reduce the indicators of inflammation caused by CRG.\nThis was evident in the decrease in the levels of these cells of all types in\nthe C&E and ECP groups, while the rest of the levels of these cells were\nhigh in the CRG group. This confirms the presence of anti-inflammatory\nsubstances in the aqueous extract of Eucalyptus<\/em> leaves61,62<\/sup>.<\/p>\n\n\n\n

CRG-induced inflammation allows for the evaluation of cell\nmigration, the involvement of cytokines and chemical mediators, and protein\nleakage. While ECP did decrease the overall leukocyte count in the peripheral\nblood, it is worth noting that neutrophils comprised the largest proportion of\nleukocytes in the group treated with CRG. We can see that this ECP can lower\nthe amounts of inflammatory substances made by immune cells because the ratio\nof neutrophils went down and the levels of monocytes, lymphocytes, CRP, and TNF\nwent down in the blood of the test animals63<\/sup>.<\/p>\n\n\n\n

The effect of ECP on CRP<\/strong><\/p>\n\n\n\n

The liver\nproduces CRP in response to inflammatory cytokines64<\/sup>, CRP levels\nrise when the liver is exposed to oxidative stress65<\/sup>, indicating\nsystemic inflammation and linked to chronic diseases like cardiovascular disorders\nand autoimmune conditions66<\/sup>. Since CRG\nenhances levels of inflammation that in turn increase CRP levels, which is what\nwas observed in the CRG group, ECP reduces the level of inflammation that in\nturn decrease CRP levels (Table 2), which is what we observed in the C&E\ngroup67<\/sup>. Eucalyptus\nglobulus<\/em> leaves’ essential oils, particularly eucalyptol, have\nanti-inflammatory properties, preventing pro-inflammatory cytokine synthesis\nand encouraging anti-inflammatory mediator release, potentially lowering CRP\nlevels68<\/sup>. Eucalyptus\nglobulus<\/em> extracts enhance antioxidant activity, reduce oxidative stress,\nlower inflammatory responses, and reduce CRP levels69<\/sup>. Essential\noils are known to possess significant antioxidant action, which is related to\nphenolic groups like thymol, carvacrol, and maybe 1,8 cineole70<\/sup>.<\/p>\n\n\n\n

The Effect of ECP on TNF<\/strong><\/p>\n\n\n\n

The cytokine\nTNF, which is generated by lymphocytes (T-cell)71<\/sup>, and monocytes\n(macrophages)72<\/sup>, plays a\ncrucial role in immune cell control and systemic inflammation73<\/sup>, triggering a\nrange of immunological responses74<\/sup>. Although\ncytokines are not the only routes or substances that cause inflammation75<\/sup>, TNF plays a\ncritical role as a mediator in the start and spread of inflammatory reactions76<\/sup>. The CRG showed higher\nTNF levels due to abnormally increased lymphocytes and monocytes, which are key\nsecretors of TNF. Conversely, a decrease these cells in C&E group resulted\nin a decrease in TNF levels in this group (Table 2). The antioxidant property of\nEucalyptus\nglobulus<\/em> <\/em> leaves is due to the following\nmonoterpenes77<\/sup>: \u03b1 pinene, \u03b2 pinene, limonene, \u03b2 myrcene, sabinene and terpinolene have\nantioxidant properties56<\/sup>.<\/p>\n\n\n\n

Table 1: Evaluation of Blood parameters in different experimental configurations<\/strong><\/p>\n\n\n\n\n\n\n\n\n\n
\n

Groups<\/strong><\/p>\n<\/td>\n

\n

WBC<\/strong><\/p>\n

(x103<\/sup>\/\u03bcL)<\/strong><\/p>\n<\/td>\n

\n

Lymphocytes<\/strong><\/p>\n

(x103<\/sup>\/\u03bcL)<\/strong><\/p>\n<\/td>\n

\n

Monocytes<\/strong><\/p>\n

(x103<\/sup>\/\u03bcL)<\/strong><\/p>\n<\/td>\n

\n

Neutrophils<\/strong><\/p>\n

(x103<\/sup>\/\u03bcL)<\/strong><\/p>\n<\/td>\n<\/tr>\n

\n

Control<\/p>\n<\/td>\n

\n

04.00 \u00b1 0.00<\/p>\n<\/td>\n

\n

07.08 \u00b1 0.15<\/p>\n<\/td>\n

\n

00.25 \u00b1 0.03<\/p>\n<\/td>\n

\n

01.55 \u00b1 0.19<\/p>\n<\/td>\n<\/tr>\n

\n

ECP<\/p>\n<\/td>\n

\n

04.85 \u00b1 0.31 a<\/sup><\/p>\n<\/td>\n

\n

07.36 \u00b1 0.20 a<\/sup><\/p>\n<\/td>\n

\n

00.20 \u00b1 0.02<\/p>\n<\/td>\n

\n

01.63 \u00b1 0.09<\/p>\n<\/td>\n<\/tr>\n

\n

CRG<\/p>\n<\/td>\n

\n

13.50 \u00b1 0.98 ab<\/sup><\/p>\n<\/td>\n

\n

11.10 \u00b1 0.13 ab<\/sup><\/p>\n<\/td>\n

\n

01.97 \u00b1 0.08 ab<\/sup><\/p>\n<\/td>\n

\n

04.77 \u00b1 0.12 ab<\/sup><\/p>\n<\/td>\n<\/tr>\n

\n

C&E<\/p>\n<\/td>\n

\n

08.83 \u00b1 0.75 ab<\/sup><\/p>\n<\/td>\n

\n

09.42 \u00b1 0.09  ab<\/sup><\/p>\n<\/td>\n

\n

01.16 \u00b1 0.08 ab<\/sup><\/p>\n<\/td>\n

\n

03.79 \u00b1 0.09 ab<\/sup><\/p>\n<\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

 <\/p>\n\n\n

The values are mean \u00b1 SD value, n = six in each group, a <\/sup>show\nthe difference in statistics. With a control group, b<\/sup> statistical\ndisparity according to illness group, (P < 0.05). ECP: Group of aqueous\nextract of Eucalyptus globulus<\/em> leaves, CRG: group of Carrageenan, C&E: group of carrageenan then aqueous extract of Eucalyptus\nglobulus<\/em> leaves.<\/p>\n\n\n\n

Table 2: Evaluation of Serum parameters in different experimental configurations<\/strong><\/p>\n\n\n\n\n\n\n\n\n\n
\n

Groups<\/strong><\/p>\n<\/td>\n

\n

CRP<\/strong><\/p>\n

Mg\/dl<\/strong><\/p>\n<\/td>\n

\n

TNF<\/strong><\/p>\n

Pg\/mL<\/strong><\/p>\n<\/td>\n<\/tr>\n

\n

Control<\/p>\n<\/td>\n

\n

0.10 \u00b1 0.10<\/p>\n<\/td>\n

\n

127.83 \u00b1 8.33<\/p>\n<\/td>\n<\/tr>\n

\n

ECP<\/p>\n<\/td>\n

\n

0.12 \u00b1 0.10<\/p>\n<\/td>\n

\n

133.50 \u00b1 3.51<\/p>\n<\/td>\n<\/tr>\n

\n

CRG<\/p>\n<\/td>\n

\n

0.60 \u00b1 0.33 ab<\/sup><\/p>\n<\/td>\n

\n

243.50 \u00b1 18.59 ab<\/sup><\/p>\n<\/td>\n<\/tr>\n

\n

C&E<\/p>\n<\/td>\n

\n

0.37 \u00b1 0.21 ab<\/sup><\/p>\n<\/td>\n

\n

186.00 \u00b1 3.85 ab<\/sup><\/p>\n<\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

 <\/p>\n\n\n

The values are mean \u00b1 SD value, n = six in each group, a <\/sup>show\nthe difference in statistics. With a control group, b<\/sup> statistical\ndisparity according to illness group, (P < 0.05). ECP: Group of aqueous\nextract of Eucalyptus globulus<\/em> leaves, CRG: group of Carrageenan, C&E: group of carrageenan then aqueous extract of Eucalyptus\nglobulus<\/em> leaves.<\/p>\n\n\n\n

Conclusion <\/strong><\/p>\n\n\n\n

A simple method similar\nto Iraqi folk medicine was used to get antioxidant and anti-inflammatory\ncompounds from Eucalyptus globulus leaves. The study found that rats exposed to\ncarrageenan for 30 and 15 days had much lower levels of inflammation than rats\nin the control group.<\/p>\n\n\n\n

The study identifies\nparameters like WBC, lymphocytes, monocytes, neutrophils, CRP, and TNF to\nestablish relationships between them. It recommends research on leaf extract’s\neffects on the respiratory system, bronchi, and bronchioles.<\/p>\n\n\n\n

Acknowledgement<\/strong><\/p>\n\n\n\n

I would like to thank the Faculty of Nursing, Faculty of pharmacology \/\nUniversity of Kerbala, Iraq.<\/p>\n\n\n\n

Funding Source<\/strong><\/p>\n\n\n\n

The authors received no\nfinancial support for the research, authorship, and\/or publication of this\narticle<\/p>\n\n\n\n

Conflict of Interest <\/strong><\/p>\n\n\n\n

The authors do not have\nany conflict of interest.<\/p>\n\n\n\n

Data Availability Statement<\/strong><\/p>\n\n\n\n

This statement does not apply to this article.<\/p>\n\n\n\n

Ethics Statement<\/strong><\/p>\n\n\n\n

This research did not involve human participants, animal subjects, or any material that requires ethical approval<\/p>\n\n\n\n

Informed Consent Statement<\/strong><\/p>\n\n\n\n

This study did not\ninvolve human participants, and therefore, informed consent was not required<\/p>\n\n\n\n

Author\u2019s contribution<\/strong><\/p>\n\n\n\n

Mustafa K. Mushatet: Conceptualization, Methodology, Writing \u2013 Original Draft.<\/p>\n\n\n\n

Asaad Abbas khalaf: Analysis, Writing \u2013 Review and Editing.<\/p>\n\n\n\n

Doaa A. Hamad: Visualization, Supervision, Project Administration.<\/p>\n\n\n\n

Thikra Abd Jary: Data Collection, Funding Acquisition, Resources, Supervision<\/p>\n\n\n\n

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