Medulloblastoma (MB), occurring in the cerebellum, is the most common childhood brain tumor. Because conventional methods decline life quality and endanger children with detrimental side effects, computer models are needed to imitate the characteristics of cancer cells and uncover effective therapeutic targets with minimum toxic effects on healthy cells. In this study, metabolic changes specific to MB were captured by the genome-scale metabolic brain model integrated with transcriptome data. To determine the roles of sphingolipid metabolism in proliferation and metastasis in the cancer cell, 79 reactions were incorporated into the MB model. The pathways employed by MB without a carbon source and the link between metastasis and the Warburg effect were examined in detail. To reveal therapeutic targets for MB, biomass-coupled reactions, the essential genes/gene products, and the antimetabolites, which might deplete the use of metabolites in cells by triggering competitive inhibition, were determined. As a result, interfering with the enzymes associated with fatty acid synthesis (FAs) and the mevalonate pathway in cholesterol synthesis, suppressing cardiolipin production, and tumor-supporting sphingolipid metabolites might be effective therapeutic approaches for MB. Moreover, decreasing the activity of succinate synthesis and GABA-catalyzing enzymes concurrently might be a promising strategy for metastatic MB. Medulloblastoma (MB) is the most common brain tumor in children with an incidence of five per million ,2. MB isMetabolic changes stemming from medulloblastoma include the over-expression of enzymes related to lipid production and the Warburg effect in which abnormal glucose consumption and overproduction of lactate are observed regardless of oxygen presence ,11. ReduFurthermore, MB cells were found to upregulate the amino acid transporter SLC1A5, which takes up glutamine in cells . High acIn the present work, the genome-scale brain metabolic model iMS570 reflects healthy brain metabolism and was constructed previously by our group  and thenThe genome-scale metabolic brain models are reconstructed for healthy and MB cases (nine MB models) and integrated with transcriptomic data (three GSE data incorporated with Gene Inactivity Moderated by Metabolism and Expression (GIMME)). Two MB models were reconstructed by using the average of all gene expression levels in GSE62600 and GSE37418 in order to compare the results with each other and with the healthy results. Four MB-specific models that reflect subtypes of MB were created and named WNT, SHH, GR3, and GR4 using the GSE37418, including gene expression levels of these subtypes. Lastly, the non-metastatic (M0) model, metastatic (M2), and (M4) MB models were developed employing GSE10327 to find the relation between the Warburg effect and metastasis. The models and simulation results were validated by comparing the FBA results with the data obtained experimentally in the literature. The main reactions carried out in the cell are presented in The substantial metabolic differences between the MB and healthy cases and the experimental findings in the literature are summarized in Despite low TCA flux rates in the MB models, acetyl-CoA formation (R28 + R72) in the TCA cycle, which is necessary for lipid synthesis, is higher compared to the healthy brain model. Acetyl-CoA is produced in the pyruvate dehydrogenase reaction (R13 + R57) and threonine metabolism (R197 + R200). The glutaminase reaction (R96) showed higher activity in all MB models than the healthy model, which is consistent with the literature where the low expression levels of glutamate uptake proteins, EAAT1 and EAAT2-4 were observed . GlutamiThe ratio of glutamate over glutamine accumulation in the cerebellum predicted by the MB model was found to be very close to the concentration ratio determined experimentally  Table 1Table 1. When glucose is available, the MB cells mainly use aerobic glycolysis because of the Warburg effect. Even though OXPHOS was also utilized by MB cells, the importance of glycolysis for both energy production and cell proliferation is undeniable. Additionally, due to the heterogeneity of the tumor environment, tumor cells may require different metabolic needs. Using FBA, the glucose uptake rate was restricted to zero to be able to observe the impacts of glucose deficiency on energy production in the MB. MB, which utilizes glycolysis regardless of oxygen availability, was observed to be almost entirely reliant on OXPHOS when glucose is not available. In this case, whole energy generation was reduced by 82% a and theThen, to simulate the effects of a glutamine deficiency, two consecutive adjustments to the MB model, glutamine uptake and both glutamine and glucose uptake rates, were done where these uptake rates were reduced to zero.In the absence of glutamine, energy generation in the TCA cycle decreased to zero. In both glutamine and glucose deficiencies, energy generated in the TCA cycle declined from 6% to 4% b. In botThen, biomass rate (R25) was also examined in cancer cells to investigate the impact of carbon source deficiency on the growth of MB cells c. It wasHexokinase 2 (HK2), Phosphofructokinase (PFK), and Pyruvate Kinase (PK) genes in metastatic samples, the ratio of ATP produced in glycolysis over ATP produced in OXPHOS (ATPG/ATPOP) was detected to be almost seven times higher in MB-M2 than the ATPG/ATPOP found in MB-M0  procedure, 205 reactions out of 594 were estimated to be significantly different from one another. In the MB neuron cells, 73% of reactions (11 out of 15 reactions) in the glycolysis pathway showed statistically important deviation from the healthy neuron cells because of the increased glucose uptake and excessive lactate production. As was observed in the FBA result, lactate production was determined to be statistically higher relative to the outcome for the healthy model  and gene expression data values (ZG) that were higher than 1.96 or lower than −1.96 were categorized as transcriptionally regulated reactions and reactions that are not regulated at the transcriptional level (NTR). The reactions where both flux and gene expression alter are called transcriptionally regulated (TR) . As a reIn glycolysis, most of the reactions are not regulated at the transcriptional level . This stIt was detected that lactate dehydrogenase in the glycolysis pathway has an increased ZF score, although its ZG score is remarkably low. A high level of pyruvate might be causing an increase in lactate formation.Fatty acid synthesis had the highest TR ratio with more than one-third of reactions being transcriptionally regulated . ACOT7 gene controls the stearate production from stearoyl-CoA and FADS2, and SCD, the genes that regulate oleoyl-CoA production from stearoyl-CoA. Due to significantly low ZF and ZG scores for the ACOT7, FADS2, and SCD genes, it is pertinent to point out that the oleoyl-CoA production is downregulated in MB and there is a decrease in oleic acid quantity in MB relative to the healthy brain. Oleoyl-CoA, which is the metabolically active form of oleic acid, has anti-cancer features [The features ,26. Basefeatures . Therefoasparagine synthetase(ASNS) gene regulating this reaction was upregulated in some cancer types such as pancreatic, ovarian, and prostate [ASNS, which was found by the MB model in both analyses, could be a potential therapeutic target for MB.The reactions related to biomass reactions that do not affect the vital functions of healthy cells are counted as potential drug targets to suppress tumor growth. The reactions controlling the uptake of essential amino acids including phenylalanine, lysine, and histidine  were fulprostate . Reducedprostate . Thus, AThe reactions associated with the production of cardiolipin, which is used in the structure of the cell membrane , are dirConsistent with other analyses performed in this work, the reaction where cholesterol produced in an astrocyte is transferred to the neuron was also found to be directly coupled to the growth reaction (R25).Single/double gene and reaction deletion analyses were performed to be able to find potential therapeutic targets for MB. In essentiality analysis, eight genes out of forty-two were removed as they significantly affected the main functions of the healthy cells see . When thThe essential reactions were found in pathways such as phosphatidylethanolamine, cholesterol, sphingomyelin, glycosphingolipid, phosphatidyl-inositol, and fatty acid metabolism. In agreement with the experimental findings in which lipid was found abundantly in MB , lipid pASNS, which is associated with several malignancies [PGS and CRLS1, and the glycerol-3-phosphate syntheses genes GPD1 and GPD2 were found as essential genes. Glycerol-3-phosphate is utilized in cardiolipin metabolism to produce cardiolipin, making up 20% of the mitochondrial membrane [GPD2 was observed to be excessively active in numerous cancers [GPD2 inhibition led to anti-cancer effects in a prostate cancer cell line [GPD1 and GPD2 genes and the reactions controlled by them were found in both flux coupling and single-gene deletion analyses by the MB model. Furthermore, inhibition of these genes did not cause severe changes in the vital functions of the healthy model.Many of the controlling genes/gene products related to the reactions detected in flux coupling analysis were also found in essentiality analyses. For instance, gnancies , was idemembrane . GPD2 wa cancers . Furtherell line . ConsistST3GAL5 gene regulating GM3 synthesis was identified as essential with the MB model and GM3 was found abundantly in MB by Chang et al. [ST3GAL5 gene was highly active in comparison to the control group and other subtypes [The g et al. . Even thg et al. ,39. Partsubtypes . Hence, B4GALNT1 gene that controls GM2, GD2, and GT2 syntheses using GM3, GD3, and GT3 gangliosides, respectively [Another essential gene found by the MB model was the ectively . The mosectively ,41,42. Tectively . Only MBectively .In double gene deletion analysis, 26 synthetic lethal gene pairs were determined once the genes found in single deletion analysis were removed . HoweverATP citrate lyase (ACL) and Fatty Acid Synthase (FASN) genes was captured by the MB model in double gene deletion analysis. In agreement with our simulations, simultaneous inhibition of the ACL and FAS genes hindered cancer cell division [ACL, which is responsible for the production of acetyl CoA in the TCA cycle, reduced the growth of lung cancer cells [FASN also caused a decrease in the hallmarks of tumors such as lipid production in various cancers such as pancreatic, ovarian, and lung tumors [The combination of the division . Moreoveer cells . Suppresg tumors . AlthougABAT gene with succinate-CoA ligase ADP-forming subunit beta (SUCLA2) and succinate-CoA ligase GDP/ADP-forming subunit alpha (SUCLG1) genes that regulate succinate synthesis (R34 + R77) from succinyl-CoA in the TCA cycle were also determined by the MB model (GABA transaminase (ABAT), which controls SuccinateSAL and glutamate productions (R99 + R101) from alpha-ketoglutarate and GABA, is relatively lower in primary MB than in healthy cerebellum since overexpression of ABAT in MB decelerates proliferation and increases the activity of OXPHOS [ABAT was found to be less active than SHH and WNT. Furthermore, activation of ABAT was observed to trigger apoptosis and diminish cancer cell division in rats with MB [ABAT expression was found to be enhanced in metastatic MB in comparison to primary MB in order to meet its energy needs by utilizing GABA [ABAT gene was found to be crucial for metastatic cells because ABAT recharges the metabolites necessary for the TCA cycle and OXPHOS by contributing to succinate and NADH productions.Combinations of the MB model . The actf OXPHOS ,47. In fSDH), which converts succinate to fumarate, were found in some malignancies as high succinate levels cause hypoxia-inducible factor 1-alpha  stabilization and HIF-alpha controls the genes triggering cell proliferation [Succinate upregulation is also associated with cancer . Abnormaferation . HoweverSGPL1 gene, which regulates reversible phosphoryl-ethanolamine synthesis from sphingosine 1-phosphate (SP1), was identified with the KDSR and DEGS2 genes .Single gene essentiality was repeated for lactate and energy productions as the objective functions to capture the genes that are crucial for three parameters  since interfering with these genes was considered to improve the therapeutic strategy. In total, 32 common essential genes were found in the three analyses. Then, these 32 essential genes were silenced one at a time from the normal brain model to identify the ones whose knockout does not affect the necessary functions of the healthy brain cells. The genes that decrease MB growth by more than 40% were determined. Taking these criteria into consideration, the essential gene list was narrowed down to eight. By using the DrugBank and GeneCard databases, 54 drugs/compounds were found for eight essential genes  was studied in numerous tumors such as colon cancer, lung cancer, ovarian cancer, melanoma, and GBM . StudiesFASN was determined to be overexpressed [Antimetabolites are chemically similar substances to natural metabolites and they are often used for cancer and viral diseases to inhibit the use of natural metabolites . To idenxpressed .HMGCR, was found to be considerably similar to mevalonate with a 0.91 Tanimoto score; however, it crosses the blood–brain barrier with poor transport capacity [Metabolites of cholesterol metabolism were found to make up 10% of the total detected substrates. These metabolites are mostly intermediates of the mevalonate pathway, which is stimulated by malicious cells to support cancer-associated features such as uncontrollable growth . In addicapacity .Fifteen percent of natural metabolites consumed in reactions whose catalyzing enzymes are potential targets are linked to sphingolipid metabolism. While 8% of these metabolites were determined to be associated with the sphingomyelin pathway, 7% of them are related to the glycosphingolipid pathway. Indeed, enzymes and intermediates associated with sphingolipid metabolism were determined to play vital roles in cell signaling mechanisms. One of the antimetabolite candidates detected similar to sphinganine metabolite is phytosphingosine with a 0.95 Tanimoto score. Sphinganine is used in the production of sphinganine-1-phosphate and dihydroceramide . PhytospDespite its anti-cancer features, its delivery might be challenging owing to its low solubility and permeability through the blood–brain barrier (BBB) ,75. The Considering the lack of studies on phytosphingosine delivery, further research using nano-engineering techniques including micelles, nano-capsules, liposomes, and emulsions seems promising.Among pediatric brain tumors, medulloblastoma (MB) is the most prevalent kind  with an annual rate of ~5 cases per one million population ,2. The tCRLS1, PGS1, GPD1, and GPD2) were determined as essential in the essentiality analyses. Notably, the CRLS1 gene was also identified as one of eight essential genes required for ATP and lactate production besides growth. In the essentiality analyses, 45 genes and 312 reactions were identified as essential. A total of 42 of them were detected in both analyses and almost 60% of the essential genes were associated with lipid production. In sphingolipid metabolism, the B4GALNT1 and ST3GAL5 genes were determined as essential by the MB model. The fact that inhibition of GM3 synthase led to anti-cancer effects in MB, and GM3 was found abundantly in MB, renders the ST3GAL5 gene a desirable target for MB treatment. The essentiality analyses for biomass, energy and lactate productions resulted in eight common genes , regarding their toxic effects on the healthy model and their potential as therapeutic targets for MB. The antimetabolites that result in competitive inhibition to natural substrates in reactions were discovered, and it was found that 32%, 15%, and 10% of natural metabolites in reactions whose enzymes can be used as therapeutic targets were related to fatty acid synthesis, sphingolipid metabolism, and cholesterol synthesis, respectively. Mevalonate–meglutol and sphinganine–phytosphingosine were identified to be very similar to each other with 0.91 and 0.95 Tanimoto scores, respectively. Phytosphingosine has anti-cancer properties and has the potential to act as an antimetabolite, but further experimental study is needed to fully understand how it enters the BBB.In an attempt to find the drug targets for MB, 158 reactions were estimated to be coupled with the growth reaction. Interfering with these reactions may be a potential therapeutic strategy for MB. Because various cancer agents are interfering with nucleotide metabolism and considering how the MB model captured nucleotide exchange reactions coupled to growth reaction, the MB model made the right predictions about cancer metabolism. In addition, reactions associated with the production of CDP diacylglycerol and glycerol-3-phosphate and main cardiolipin production reactions stand out in the flux coupling analysis. Moreover, cardiolipin and glycerol-3-phosphate synthesis reactions and the genes regulating them , FASN, Acetyl-CoA Carboxylase 1 (ACACA), Glutaminase (GLS) were the upper limit for the threshold value since these genes are upregulated in MB [FASN expression value and high enough to create a model which reflects the MB characteristics.The expression levels of ed in MB ,85,90,91To generate specific models for MB subtypes, four different thresholds were created by using averages of gene expression of patients in different subgroups. Taking into consideration the criteria determined, thresholds were chosen as 72.5%, 74.7 %, 90.24%, and 76.34% of the average of WNT, SHH, GR3, and GR4 gene expressions, respectively. GIMME deleted 7, 7, 9, 13, and 8 reactions from MB, WNT, SHH, GR3, and GR4 models, respectively.GLS expression value. As in GSE37418, the expression levels of upregulated genes in MB were determined as an upper bound. Consequently, 30% of the transcriptome data values average was selected, and 38 reactions were erased by GIMME from the MB model for GSE62600.The differences between gene expression values in GSE62600 were higher compared to GSE37418. Thus, small variations in the threshold did not influence the results remarkably. The thresholds that are more than 50% of the transcriptome data average of MB were eliminated because these values were exceeding the To investigate the correlation between metastasis and the Warburg effect, the expression values of M0, M2, and M4 medulloblastoma metastasis stages that are present in GSE10327 were utilized to construct MB models that reflect metastasis-specific properties. As thresholds, 45%, 50%, and 55% of the gene expression averages were selected for the non-metastatic (M0), M2, and M4 stages, respectively. GIMME eliminated 31, 31, and 26 reactions from the non-metastatic (M0) model, metastatic (M2), and (M4) MB models, respectively.The context-specific constraints imposed on the system include the uptake of glucose, glutamine, oxygen, glycogen, ketone bodies, leucine, tyrosine, tryptophan, methionine, and ammonia. Since similar proliferative propensities were observed for both glioblastoma (GBM) and medulloblastoma (MB) cell lines, in 5 mL of glutamine for 10 days , 0.080 mThe tryptophan and tyrosine uptake rates of MB over the ones in the healthy cerebellum were determined as 3.5–3.7 and 2.8, respectively ,94. UsinAll simulations were carried out using CPLEX ILOG optimization algorithms provided by IBM’s Academic Initiative Program on the COBRA toolbox under MATLAB 2017b. The objective function was set as maximization of biomass reaction if not otherwise stated, and the flux balance analysis was conducted after the integration of transcriptome data. Next, the minimization of the metabolic adjustment method (MOMA) was performed to predict closer suboptimal results to the healthy case in order to improve the flux results acquired with FBA ,96. The The objective function for MOMA was determined as the minimization of the distance (D) between medulloblastoma (m) and healthy (h) cases. For medulloblastoma (m) and healthy (h) cases, flux rates obtained from FBA in MB and healthy (iMS570) models were utilized. Since this expression includes quadratic terms, quadprog function found in optimization toolbox of MATLAB was employed.Flux sampling method was conducted by employing the gpSampler function of the COBRA Toolbox. This function uses Artificially Centered Hit-and-Run (ACHR) algorithm that estimates the random flux profiles . In totaFlux sampling results were also used for the regulation analysis that incorporates flux data with gene expression data by transforming flux and DNA microarray data into statistical scores. Z scores for metabolic fluxes (ZF) and gene expression data values (ZG) were estimated to find the significance of change between MB and healthy conditions.The reactions whose flux values and/or expression levels of the genes controlling these reactions were higher/lower than 1.96 and −1.96 which were computed with respect to a 5% significance level, were categorized regarding whether they are metabolically or transcriptionally, or post-transcriptionally controlled. ZG scores of some reactions whose controlling genes are more than one were estimated by summing up all ZG scores designated for each gene.Flux coupling analysis (FCA) was executed with the F2C2 function to uncover reactions that have a direct impact on biomass reactions. Hence, the reactions that can be intervened to hinder growth in MB were revealed. Then, these potential therapeutic targets, which have no destructive impact on healthy cells, were detected.The essentiality analyses were conducted where all reactions or genes are erased one at a time from the model. If the removal of components leads the flux rate of growth reaction to decline less than the specified flux rate (40%), they are acknowledged as essential. The genes required for energy generation and lactate production along with biomass production rate for the survival of tumor cells were uncovered. Then, the reactions regulated commonly by these essential genes identified in three analyses  were deleted one at a time from the healthy brain and MB models and the genes controlling eliminated reactions that do not influence the basic functions of the healthy cells and decrease the MB growth rate by more than 40% were estimated.To predict the compounds targeting metabolic enzymes and their impact on both medulloblastoma and healthy cells, the antimetabolites  were explored by using Tanimoto scores and performing FBA. Antimetabolites might diminish the usage of substrates in cells by engendering competitive inhibition; therefore, all the compounds similar to the metabolites present in the medulloblastoma brain model were determined from DrugBank using Simplified molecular-input line-entry system (SMILES) of these metabolites and the compounds with Tanimoto scores higher than 90% were identified . Then, t


Low-grade gliomas are the most common brain tumors in children. This tumor type presents a wide range of clinical, histological, and biological behaviors. In recent years, an association between estrogens and progesterone and the development of tumors has been suggested. A case of a 2-year-old girl is described with a low-grade brain tumor treated with chemotherapy and disease stabilization. The treatment with Decapeptyl® was initiated due to precocious puberty, and the tumor showed a decrease in its solid component—more than 50% of the initial size—three years after starting treatment. Several studies have described the influence of estrogen and progesterone on the development of gliomas, decreasing or increasing their expression in those tumors with greater aggressiveness, respectively. Despite the fact that the tumor-hormonal expression relationship in other tumor types has been evaluated, its role in the treatment of brain tumors remains unknown. Brain tumors are the second most common cause of childhood cancer in children between 0 and 14 years of age (approximately 22%) and the third most common cause of cancer in adolescents between 15 and 19 years of age  .The LGG constitute a heterogeneous group of tumors regarding clinical symptoms, histology, and biological behavior. Up to 5–10% can present metastases in other locations at diagnosis . The pilThe annual incidence of LGG is close to 10–12 cases per million children under fifteen years of age, with a male-to-female ratio of 1.1:1/1.2:1. Although the symptoms may vary, most are indolent and do not transform into malignant tumors (even cases of spontaneous regression have been described) . On the Estrogens are steroid hormones that affect different systems: reproductive, gastrointestinal, skeletal, immune, and CNS. Most of its effects are mediated by alpha (ER-α) and beta (ER-β) estrogen receptors ,9. SeverThis manuscript describes our experience with a pediatric patient with an LGG and her response to gonadotropin-releasing hormone (GnRH) analog drugs.2/dose) was initiated for 70 weeks, according to the recommendations of “Vinblastine in recurring or refractory pediatric low-grade glioma” [In November 2017, a two-year-old boy and a one-month-old girl were consulted in the Pediatric Emergency Department of a tertiary-level hospital due to persistent vomiting, gait disturbances, and noticeable instability after one week of evolution. Given this clinical picture, a head computer tomography was performed, revealing a mass in the supratentorial region with a significant component of hydrocephalus. The study was completed with cranial magnetic resonance imaging (MRI) (a single mass located in the suprasellar region with a solid and cystic component was found). The hormonal studies , tumor markers (AFP and B-HCG), and visual and auditory potentials  were conducted. A ventriculoperitoneal (VP) shunt was placed to improve the symptoms of intracranial hypertension, and a biopsy of the lesion was performed during the same surgical act. A pathological report confirmed the etiology of pilomyxoid astrocytoma  with a KIAA1549-BRAF mutation. In December 2017, chemotherapy based on weekly Vinblastine . The tuIn the control MRI performed two months after starting treatment with triptorelin, a decrease in the solid component of the tumor was observed, along with a significant decrease in contrast uptake. In successive controls, this decrease resulted in more than 50% less compared to the initial size without contrast uptake  . The PatRegarding hormonal controls, IGF-1 and IGFBP3 values showed a progressive increase, with an altered response to repeated oral glucose overload tests. Given the suspicion of failure in the growth hormone axis that could explain the non-control of growth in our patient with the use of triptorelin, treatment with Octreotide acetate, 1/2 vial i.m., was started in September 2021 every 28 days, being able to space triptorelin every ten weeks. Latest anthropometric data and physical examination at seven years and five months: Weight: 32.5 kg (+1.15 SD), Height: 138.6 cm (+2.66 SD), Growth rate: 4.7 cm/year , BP 97/65 (p30/p63). Thelarche 1, not Axilarche ubic hair on the labia majora, not on the mount of Venus.The steroid hormones have a prominent neuroprotective role in neurological disorders, such as Parkinson’s or Alzheimer’s disease , due to their antioxidant properties or activation . HoweverSeveral studies have described the influence of estrogens on the development of gliomas through the direct interaction with their intracellular receptors ER-α and ER-β , 18,19 . In addition, although the effect of chemotherapy in reducing the tumor size can be observed sometime after its initiation, in our case the relationship with the observed response more than one year after the treatment initiation without dose reduction is improbable. Moreover, some cases of spontaneous regression in children with low-grade gliomas have been reported, in contrast to adult patients who tend to show more aggressive behavior ,38. In oThe treatment with triptorelin in this patient led to an evident reduction in the size of the tumor that was not achieved with chemotherapeutic treatment. Moreover, this treatment had no side effects and was well tolerated. To date, this is a promising response, although the outcome after the completion of therapy is unknown. Thus, more extensive studies in the pediatric population are needed to obtain more reliable data on the effectiveness of hormonal treatment for this type of tumor, assess the situation once treatment is discontinued, and also evaluate the development of long-term side effects.


TYR, OCA2, TYRP1, SLC45A2, SLC24A5, C10orf11, and DCT) and one locus (OCA5) in consanguineous and sporadic albinism. The complications of OCA result in skin cancer and variable syndromes such as Hermansky–Pudlak syndrome (HPS) Chediak–Higashi syndrome (CHS). In the Pakistani population, autosomal recessive non-syndromic OCA is common and is associated with a large number of consanguineous families, and mutations in genes of non-syndromic types are reported. This review highlights the updates on the genetic mutation of OCA genes reported from Pakistani families. Several studies reported the genetic mutations in OCA1, OCA2, OCA3, OCA4, and OCA6 albinism in Pakistani families. A locus, OCA5, was also reported from the Pakistani population, but the gene has not been identified. A new type of OCA8 was identified due to the DCT gene mutation, and it is also reviewed here.Oculocutaneous albinism (OCA) is an autosomal recessive syndromic and non-syndromic defect with deficient or a complete lack of the melanin pigment. The characteristics of OCA appears in skin, hair, and eyes with variable degree of pigmentation. Clinical manifestations of OCA include nystagmus, photophobia, reduced visual acuity, hypo-plastic macula, and iris trans-illumination. There are eight OCA types (OCA1–8) documented with non-syndromic characteristics. Molecular studies identified seven genes linked to the OCA phenotype ( Albinism is a complex group of disorders that lead to a reduced production or complete lack of pigment–protein melanin, resulting in a characteristic white or albino appearance of hair, skin, and eyes. There are two forms of albinism, categorized as ocular (only pertaining to eyes) and oculocutaneous albinism . In these classes, the most commonly distributed entity is OCA that is reported from different ethnic, language, and ancestral groups worldwide. Clinical characteristics of OCA are associated with dramatic effects on skin, hair, and eyes. Clinical presentation of skin is widely diverse and linked with a degree of severity contingent on OCA sub-clinical types. The ocular arrangements depend on the signaling for melanin during the uterine development of the eye; hence, diverted optic nerve fibers develop ocular manifestations of the related defects ,2,3.At present, there are eight clinical types of non-syndromic oculocutaneous albinism (OCA1–8) that are linked to variable clinical features pertaining to skin, hair, and eyes. In non-syndromic conditions, each clinical type is linked to a specific causative gene. On the other hand, syndromic albinism is comprised of a group of signs and symptoms including nystagmus, visual acuity, foveal hypoplasia, and bleeding defects. At present, there are different classes of syndromic albinism in which genetic mutations are documented. There is extensive literature that describes the syndromic forms of albinism including Hermansky–Pudlak syndrome (HPS) and Chediak–Higashi syndrome (CHS). Among syndromic forms, HPS is the most common variant, and eleven subtypes (HPS1–11) are documented. The lack of ocular and dermal pigment in syndromic types of albinism is comparable to non-syndromic albinism. However, the genes impede syndromic forms that code the proteins that are extensively involved in cellular function. In syndromic albinism, the loss of function mutations produces predictable consequences. In HPS, the bleeding diathesis causes variation in genes that encode lysosomal synthesis (not only melanin synthesis involvement) and infection progression in CHS. There are other anomalies that present similar to albinism including congenital nystagmus and generalized hypopigmentation. The examples of syndromes associated with similar mutated genes as in the OCA2 type are Angelman (AS) and Prader–Willi (PWS) syndromes ,5,6.OCA is a genetic disorder with an autosomal recessive mode of inheritance in which there is a complete absence or decreased production of melanin that causes hypo-pigmentation of hair, skin, and eyes. Each parent transfers a copy of the faulty gene to the affected child in OCA. Physically normal carriers of OCA can transfer the disease to the next generation due to the presence of only one copy of the defective gene or compound heterozygous mutations in two different alleles of OCA genes, which have also been identified in different populations of the world, establishing the heterogeneity of this disorder .Albinism is also present in syndromic form with variable signs and symptoms such as neurological problems, with high susceptible to infections . HermansOn the basis of genetic testing, non-syndromic OCA is further divided into eight subtypes (OCA1–8), among which OCA1 is the most common that affects almost 50% of albino individuals worldwide .TYR, OCA2, TYRP1, SLC45A2, SLC24A5, C10orf11, and DCT) in modulating the regulation of melanin production [l-tyrosine into L-DOPA, and then dopaquinone is produced. Apart from tyrosinase a, some proteins present in melanosomes, including tyrosinase-related protein types 1 and 2 , have key functions in eumelanin production. The role of TYRP1 is to increase the ratio of eumelanin: pheomelanin production, while it also plays its role as an anti-oxidant due to the presence of peroxidase activity [TYR, OCA2, TYRP1, TYRP2, SLC24A5, DCT, MATP, and MC1R, which contribute to phenotypes of subtypes of OCA from OCA1–8 with an autosomal recessive non-syndromic pattern [In melanosomes, the molecular mechanism regulates the synthesis of melanin pigment by melanocyte cells. In this pathway, there are a number of genes and contributing enzymes that take part in multiple reactions to make melanin products. In a chemical reaction, L-tyrosine is converted into a 3,4-dihydroxyphenylalanine (DOPA) intermediate that then results in the formation of dopaquinone. From dopaquinone, the melanin synthetic pathway splits into two pathways leading to eumelanin formation (brownish to black coloration) and pheomelanin (reddish to yellow pigmentation) ultimately . Previouactivity ,13. A nu pattern .Overall, the prevalence of OCA is projected at a frequency of 1 in 17,000–20,000. The mutant allele of OCA2 is carried in approximately 1 in 70 individuals, categorized as the prevalent type of albinism worldwide, although the frequency of mutation detection is higher in the OCA1 phenotype. The frequency of OCA2 alleles is more prevalent in the sub-Saharan African population, with a rate of 1 in 1000 cases ,16. ThisClinically, there are seven OCA forms that present a wide range of phenotypes, signs, and symptoms. OCA1 is the frequent type that is further divided into OCA1A and OCA1B subtypes ,28,29,30During the embryonic development stage, the melanin deficiency leads to eye developmental defects such as the retina and iris lacking melanin pigment, foveal hypoplasia, misrouting of optic nerve fibers of the visual cortex, and small or elongated globes . As a reClinical characteristics of affected individuals are heterogeneous. OCA1A (MIM# 203100) is the subtype of OCA1, with white hair, eyebrows, eyelashes, and skin. There is no change in color throughout life. The iris color appears light-blue to pink and is absolutely transparent. The outcome of optic nerve misrouting is intense photophobia and nystagmus, and amelanotic nevi may be present. The cases with OCA1A show visual acuity 1/10 or even less, which persists throughout life without any difference between race and ethnicity .Another subtype of OCA is OCA1B (MIM# 606952), which was previously known as yellow albinism. This type is similar to the OCA1A clinical characteristics, but there is scarce development of melanin pigment in the skin and hair. This pigmentation increases with age, and the iris color may be transformed to green/brown from blue color that is present at birth. Visual acuity is 2/10 in the OCA1B type. Another variant of OCA1 is temperature-sensitive OCA in which initially non-pigmented body hair is present but pigmented hair of hands and feet may develop with time due to temperatures difference between exposed and covered body parts ,36.OCA2-affected individuals may have some melanin pigmentation that imparts a wide range of skin, hair, and eye color; however, the color usually remains lighter in comparison to the relatives of the family member who are unaffected. Hair color is usually not completely white. The OCA2-affected individuals can develop benign lesions or dark spots due to extensive sun exposure and develop moles or pigmented nevi ,16,37,38OCA3 is rufous albinism , and it was first documented in the African population. Individuals affected with OCA3 have yellow or reddish hair, red to brown skin, and brown eyes (contain excess melanin) or hazel eyes (green eyes due to less melanin). This type of OCA is reported mostly in Asian populations including Indian, Chinese, and Japanese, but is also found in Caucasian and northern European regions. In Asian origin, the color of affected individuals is fair or golden-haired, with light-brown eyebrows, and skin pigmentation is lighter compared to the skin color of the parents. In these affected albinos, the melanin pigmentation increases with age development. The degree of visual acuity impairment is less severe as examined in the affected cases of OCA1 and OCA2. Other important features of albinism including nystagmus and photophobia can be evident or may be absent ,40,41.Clinical features of OCA4 are consistent with the OCA2 features. In OCA4 cases, the range of pigmentation in the tissues varies from brown to yellowish brown. In these cases, the visual acuity can vary in severity from 20/30 to 20/400, which corresponds to the production of melanin and appears in tissue. Usually, the degree of visual acuity ranges between of 20/100 to 20/200 in OCA4 cases. OCA4 was initially documented in a case of Turkish origin but it has also expanded to other populations including German, Asian populations (Korean and Japanese), and subcontinent regions ,42,43,44OCA5 type is only reported in affected individuals of Pakistan origin. These affected family members have white skin, golden hair, and ocular impairments as seen in the OCA1 type. The amount of visual acuity was 6/60 in one affected individual of a family .OCA6 is characterized by hypopigmentation with white skin, brownish irises, and a range of hair color; golden, light brown to dark brown. Clinical features of OCA6 are not well determined because very few cases are investigated worldwide that cannot be differentiated based on specific features. The visual acuity of one affected individual measured 20/100, and fundoscopic analysis showed immature macula and revealed hypo-pigmentation. There was an absence of central fovea in the macula when compared to the brother in the same family by using optical coherence tomography . In a reThe characteristics of clinical presentations of OCA7 include skin hypo-pigmentation, and the hair coloration varies from white-golden blond to dark brown hair in affected children from normal and carrier siblings or parents. The visual acuity was reduced in the affected individuals, ranging from 6/18 to 3/60, and other ocular problems associated with iris trans-illumination and nystagmus were also observed .OCA8 is a recently investigated albinism type in Turkish and French ethnic groups. This type is a milder phenotype of albinism with moderate features including hypopigmentation of hair and skin, moderate foveal hypoplasia, nystagmus, and iris illumination . On the TYR and OCA2, and the summary of the function of OCA genes is outlined  reported to date, and only seven genes are associated with the phenotypes, while one locus is still awaiting gene identification. Of these seven genes, the most commonly mutated genes are outlined . The detTYR (MIM# 606933). The physical localization of the TYR gene is present on chromosome 11q14.3, which has five coding exons. It is involved in the regulation of biosynthesis of the tyrosinase enzyme that mediates the rate-limiting chemical reactions in melanin formation. These crucial reactions are oxidation and hydroxylation of L-DOPA and DOPAquinone, the intermediate product [TYR mutations due to defective tyrosinase activity lead to a diminished or absolute deficiency of melanin, with variable coloration in hair, skin, and eyes [TYR. On the other hand, OCA1B (MIM# 606952) is a subtype with positive tyrosinase activity that is a mild to moderate type of OCA outcomes due to less severe TYR mutations. This type of OCA presents reduced disease impact, as melanin pigmentation with less concentration is seen [The most frequent albinism is OCA1 caused by a mutation in  product . TYR mutand eyes . OCA1A  that was earlier reported as the P gene. OCA2 is physically located on the chromosome at position 15q11.2–q12 and contains 23 coding exons with 345 kb nucleotide sequences. This gene mediates a 110 kDa protein that encodes 838 amino acids and has 12 transmembrane helices [OCA2 mutations, and it is categorized as the second most putative factor for OCA development [OCA2 is the second most investigated albinism type due to the defective  helices . The P p helices . Althoug helices , it appe helices . The OCA helices ,70. In telopment ,61,63,64TYRP1 mutations (MIM# 115501). This gene spans a nucleotide sequence of 17 kb and is located on chromosome 9p23. It contains eight coding exons and encodes 536 amino acids that constitute tyrosinase-related protein-1 (Tyrp1) of ~61 kDa size. As TYRP1 has structural and conformation similarities to tyrosinase (GenBankNM_000550), it contributes to melanin synthesis by stabilizing tyrosinase and melanosome integrity and takes part in cellular proliferation and apoptosis to maintain the environment. It is also involved in the oxidation of 5,6-dihydroxyindole-2-carboxylic acid (DHICA), a step in the melanin biosynthetic pathway. Earlier studies reported OCA3-associated cases in the African population only; nucleotide variations in TYRP1 have also been investigated as follows: Caucasian origin, Asian–Indian populations, and Chinese races [The third type of albinism, OCA3 (MIM 203290), is associated with se races ,61,64,71SLC45A2 gene  gene). It is localized on chromosome 5p13, spanning a nucleotide size of 40 kb and having seven coding exons [MATP/SLC45A2 mutations were identified in Turkish ancestry, and then the reports were expanded to other populations such as the Chinese, German, Japanese, Indo–Pak, and Korean ethnic groups. This type of OCA is commonly found in the Japanese population; however, SLC45A2 mutations are rare in Pakistani OCA patients [The OCA type 4  results due to causative mutations in the ng exons . It regung exons . The pripatients ,52,55.The OCA5 locus on chromosome 4 (4q24) consists of fourteen genes, but the particular causative gene of OCA5 has not yet been identified. Kausar et al., 2013 reported OCA5 in a single Pakistani family .SLC24A5  mutations, and the gene is localized on chromosome 15q21.1 [SLC24A5 mutations were earlier identified in the Chinese population, but reports from various ethnic origins are documented that present the heterogeneity of OCA6. The manifestation of the cutaneous phenotype was different to hair color ranging from white to blond or dark brown. The SLC24A5 protein existence in melanosomes play a vital role in melanosome structure and synthesis. Therefore, a defective gene or truncated SLC24A5 protein may negatively impact the development of melanosomes and the production of melanin [SLC24A5 mutations. In Iranian families, mutations are identified as causative of OCA6 [The OCA6 type of albinism occurs due to  15q21.1 . This ge 15q21.1 . SLC24A5 melanin ,73,74. I of OCA6 .C10orf11 mutations, and this gene is physically localized on chromosome 10 at the 10q22.2–q22.3 position. C10orf11 mediates the synthesis of a Leucine-Rich Melanocyte Differentiation-Associated Protein, consisting of 226 amino acids with one LRR C-terminal (LRRCT) and three domains of leucine-rich repeats (LRRs). The C10orf11 protein plays a role in the melanin pathway including cell-adhesion, differentiation of melanocyte differentiation, assembly of the extracellular matrix, cell-signaling, RNA processing, and neuronal development [C10orf11 associated with OCA7 have been identified in Indian and Iranian populations [The OCA7 (MIM: 615179) type of albinism is associated with elopment ,75. The ulations ,76.DCT mutations investigated recently. The gene is localized on chromosome 13q31–q32 [DCT) gene, also called TYRP2, has eight coding exons and it mediates the TYRP2 protein that has 80% similarity to TYRP1. The DCT protein along with TYRP1 and TYR plays an important role in melanin biosynthesis and melanosome apoptosis [DCT have been detected in the French and Turkish populations [A new type of OCA is OCA8 (MIM 191275) linked to 3q31–q32 . The doppoptosis . Recentlulations ,48.Pakistan is situated in the subcontinent with a historical cross-side associated with the various political and cultural impeders and had been a bridge for many invaders and reigns in South Asia. Historical migrations in this region have ensued in a combination of antique cultures, which continue the social or cultural blend of this population. Inbred and close familial relationships make Pakistan a productive country to conduct genetic differentiation studies and explore the various genomic characteristics of a human population . GeneticTYR and OCA2. More than ninety consanguineous OCA families have been explored to determine the TYR mutations [TYR gene, a variety of novel, known, and founder mutations have been demonstrated by various research reports. In the population, the most frequent and common mutations of the TYR gene are c.832C>T and c.1255G>A; p.Gly419Arg that are reported in 21 families (22.6%) and 20 families (23.8%), respectively. On the other hand, some mutations are founders that contribute repeatedly to the ancestry i.e., c.230G>A (5.9%). In Pakistani consanguineous families, about 90 mutations are associated with TYR-linked OCA that includes missense 86.9%, nonsense 7.1%, and the remaining splice site error, deletion, and frameshift mutations  and the Punjabi, Sindhi, Balochi, Pashtoon, and Urdu speaking language groups. Different investigators detected genetic mutations in OCA-associated genes, predominantly utations ,63,64,65utations .OCA2. This gene is linked to more than fifty OCA families with variable mutation patterns  techniques are versatile tools to explore the hidden molecular factors associated with albinism. Using these state-of-the-art technologies might improve the diagnostic prospective and clinical understanding of the disorders. The establishment of NGS in developing countries such as Pakistan would improve the diagnostic facilities that might have an impact on disease-controlling factors. Updating the data about OCA will be helpful for researchers and clinicians to learn more about the molecular diagnostic facilities and clinical phenotypes that can improve counselling services for the families and their tailored management. Further, it might help the research to explore new ideas to determine the role of genes in therapeutic medicine. The study is limited with respect to discussing the genotype and phenotype outcome of the mutations in respective gene/s that further require comprehensive data to depict the role of these genes in the disease.Autosomal recessive non-syndromic oculocutaneous albinism (OCA) is increasingly devastating in populations with variable languages or ethnic groups, having customs of consanguineous marriages, including in Pakistan. These congenital or familial genetic disorders have augmented the health and social issues in the communities. OCA is a rare genetic anomaly with diverse heterogeneity and its clinical characteristics are evident in skin, eyes, and hair. The defects in enzymes that regulate melanin biosynthesis in melanosomes are structurally or functionally altered, and the outcome is various types of OCA. Molecular studies identified eight different loci of OCA in which seven OCA phenotypes were linked to putative gene mutations including


Nonalcoholic fatty liver disease (NAFLD) is the commonest chronic liver disease and affects a considerable proportion of the general population worldwide. Obesity is a major risk factor for development and progression of NAFLD and weight loss is an effective intervention for the management of NAFLD. However, few patients achieve substantial and sustained weight loss with lifestyle measures. Therefore, antiobesity agents are frequently considered in patients with NAFLD but there are limited data on their safety and efficacy. In the present review, we discuss the role of antiobesity agents in the management of NAFLD. All approved antiobesity agents appear to reduce transaminase levels and to improve steatosis in patients with NAFLD. However, their effects on fibrosis are less well studied and whether they affect liver-related outcomes, including progression to cirrhosis and hepatocellular cancer, is unknown. The glucagon-like peptide-1 receptor agonists, liraglutide and semaglutide, appear to represent a first-line option in obese patients with NAFLD and type 2 diabetes mellitus (T2DM) since they induce considerable weight loss and have been extensively studied in patients with T2DM. However, more studies are needed to evaluated their effects on liver-related and cardiovascular outcomes in patients with NAFLD, particularly in those without T2DM. Nonalcoholic fatty liver disease (NAFLD) is one of the most common causes of chronic liver disease. It is estimated that about 25% of the world population has NAFLD. In the United States, it is predicted that NAFLD will affect 100 million subjects by 2030. Lately, the disease is also being observed in an increased percentage of adolescents, with an estimated prevalence of 18%  , 3. Impo2 and patients with BMI 27-30 kg/m2 and at least one obesity-related comorbidity, including hypertension, diabetes mellitus (DM), dyslipidemia, obstructive sleep apnea and established cardiovascular disease.In the present review, we discuss the safety and efficacy of the available antiobesity agents in patients with NAFLD. All reviewed agents are approved for the management of patients with a body mass index (BMI) ≥ 30 kg/mOrlistat is a gastrointestinal reversible lipase inhibitor. Orlistat forms a covalent bond with the active serine residue site of gastric and pancreatic lipases resulting in inhibition of hydrolysis of dietary fats into absorbable free fatty acids and monoglycerides. Orlistat reduces dietary fat absorption by approximately 30% . Weight Several studies evaluated the role of orlistat in the management of overweight or obese patients with NAFLD. In a pilot, uncontrolled study in 10 obese patients with biopsy-proven NASH, orlistat improved steatosis in 6 patients and fibrosis in 3 patients and also reduced transaminase levels . In anotLiraglutide is a glucagon-like peptide-1 (GLP-1) receptor agonist that induces weight loss by reducing appetite and energy intake . LiragluIn a pilot, uncontrolled, 24-week study in 19 patients with biopsy-proven NASH, liraglutide reduced transaminase levels. In addition, hepatic inflammation improved in 6 patients among 10 patients who received liraglutide for 96 weeks and underwent a repeat liver biopsy at the end of treatment . In anotA number of studies evaluated the effects of liraglutide in patients with both NAFLD and type 2 DM. In an uncontrolled study (n = 68), treatment with liraglutide 1.2 mg/day for 6 months reduced body weight, visceral fat, transaminase levels as well as liver fat content measured with magnetic resonance spectroscopy . The redSemaglutide is a novel GLP-1 receptor agonist that appears to induce greater weight loss than liraglutide and has the additional advantage that it is administered once weekly, which might improve adherence to treatment . Moreovepost-hoc analysis of 2 randomized, double-blind study in obese patients with or without type 2 DM, treatment with semaglutide 0.5-1.0 mg/week for 52-102 weeks also reduced transaminase levels, which were normalized in 25-46% of the patients  neurons in the arcuate nucleus, inducing the release of α-melanocortin-stimulating hormone, which in turn acts on melanocortin-4 receptors in the paraventricular nucleus to suppress appetite , 40. Thepost-hoc analysis of 4 randomized, 56-week studies in 2073 obese patients, naltrexone/bupropion reduced transaminase levels and the FIB-4 index and this effect was more pronounced in patients with elevated transaminase levels (The combination of naltrexone, a non-selective opioid receptor antagonist, and buproprion, an inhibitor of dopamine and norepinephrine transporters, at the dose of 16/180 mg twice daily, induces weight loss approximately 4.1% greater than placebo . The mose levels . Naltrexe levels .The combination of phentermine, a sympathomimetic amine anorectic that stimulates the release of norepinephrine and epinephrine, and phentermine, an inhibitor of the norepinephrine transporter that suppresses appetite by activation of POMC arcuate nucleus neurons, at the dose of 15/92 mg/day, induces weight loss approximately 8.8 kg greater than placebo . The comSeveral agents are currently approved for the management of obesity. Most of them have been evaluated in patients with NAFLD and appear to reduce transaminase levels and to improve steatosis Table 1.AT, GN, EK and SV wrote the first draft. EC and KT revised the draft. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Plasma flow and particle interactions were exemplified in terms of momentum, energy, and turbulence flow. The transport of nanoparticles through convection, diffusion, and thermophoresis were also considered. The trajectories and heat transfer of both plasma jet fields, and particles are represented. The swirling flow controls the plasma jet and highly affects the dispersion of the nanoparticles. We demonstrate a decrease in both particles’ velocity and temperature distribution at a higher carrier gas injection velocity. The increase in the particle size and number affects the momentum transfer, turbulence modulation, and energy of particles, and also reduces plasma jet parameters. On the other hand, the upstream flame significantly impacts the particle’s behavior under velocity and heat transfer variation. Our findings open the door for examining thermal plasma impact in nanoparticle synthesis, where it plays a major role in optimizing the growth parameters, ensuring high quality with a low-cost technique.A three-dimensional numerical modelling of a time-dependent, turbulent thermal plasma jet was developed to synthetize silicon nanopowder. Computational fluid dynamics and particle models were employed via COMSOL Multiphysics Nanomaterial particles attracted immense interest in recent decades, owing to their unique characteristics. In particular, the optical, electronic, and catalytic properties change with the matter size, and differ significantly in nanometric scale to those observed in micrometer-sized particles and bulk materials ,2,3. NanAmorphous silicon nanostructures attracted considerable interest for many years, and were extensively used in several domains, such as optoelectronics, photovoltaic, and energy storage ,9,10. InVarious techniques were developed to grow silicon nanomaterials, including chemical vapor deposition, non-thermal or thermal plasma-assisted chemical vapor deposition, and laser ablation ,13,14. HThermal plasma offers a promising method for developing high-quality nanopowder with narrow size distributions ,17. Its Recently, thermal plasma achieved high-throughput fabrication of silicon nanoparticles used for lithium-ion battery electrodes. Indeed, Kambara et al.  produce In this regard, the transport of particles requires high control, as well as plasma gas composition, to ensure nanomaterials synthesis with predetermined stoichiometry . HoweverHerein, a three-dimensional numerical simulation of nanopowder formation using thermal plasma jet was provided. The velocity of the turbulent flow and the field temperature, as well as the swirl velocity effect on plasma characteristic fields, are examined. Furthermore, several process conditions, such as carrier gas velocity, size, and number of particles, and initial plasma jet properties were optimized. The effect of the variation in swirl flow intensity on the particle’s dispersion was investigated. The studied parameters reveal a high dependency on the agglomeration degree of the synthesized nanoparticles.The plasma reactor system employed for nanopowder growth contains a downward, non-transferred thermal plasma torch, as we reported in our previous work . The cat2, H2, He, or even a gas mixture. Oxidizing gases are not recommended for such a torch type, where an oxide layer may appear and covert the anode. This reduces the thermal and electric conductivity of the anode, and destruct the electrode [Several inert and reductive gases are used in DC torches, such as Ar, Nlectrode . Howeverlectrode . In DC tlectrode . The opeThe expanded argon plasma flame or jet is emitted from the plasma torch in the reactor chamber confined by the cooled cylindrical surface. Crystallized silicon particles, around 20 µm in diameter, are added to the high temperature plasma flame through argon carrier gas where they are immediately vaporized. The vapor is then transferred into the rim or tail of the plasma flame and quenched, where the temperature decreases considerably, due to the thermal expansion induced by the high-velocity flow. The silicon vapor becomes supersaturated by the rapid quenching action, and nucleates to form silicon nuclei. Simultaneously, numerous nuclei grow by heterogeneous condensation and coagulation. This coagulation process is vital for the collective development of particles. Consequently, nano-sized particles are generated . The nanThe thermal argon plasma jet injected by the torch into an argon environment reactor chamber at atmospheric pressure is represented in A 3D approach is required in this study, because when particles are injected transversely into the plasma jet, they are assumed to swirl and/or disperse in turbulent ways. Consequently, they take on 3D trajectories that need to be analyzed in 3D geometry, in order to improve accuracy of the numerical model.The 3D time-dependent model used in our study could be described by admitting the below assumptions : (a) plaTherefore, the following equations are used to simulate thermal plasma jet flow :Equation of mass conservation:Equation of momentum:u, tr) indicates the transposition.Equation of energy:The given by ,37,38:Turbulent kinetic energy Turbulent dissipation rate The production term The coefficients ε = 1.3, and σk = 1 [The considered values of turbulent constants are: d σk = 1 . The thed σk = 1 . The boundary conditions employed in this model is summarized as follows:At the jet inlet, , we admitted the assigned polynomials expression used by M. Shigeta , based oA comparison between the numerical calculation, the polynomial expression, and experimental results of axial velocity The radial velocity at the nozzle exit is not considered, In addition, the flow being swirling at the jet inlet and the azimuthal velocity are defined as :(15)uθ={The non-slip condition  we The evolution of particles position, velocity, and temperature in the plasma jet is simulated, using Lagrangian equations . Therefop, px, The particle’s position is given as:The drag force is defined as:y vector :(20)τp=4essed as :(22)CD=2The function The collisions between particles themselves, as well as with the injector wall, disperse the particle jet at the injector exit. Therefore, the fluid velocity applied for the drag force while turbulent dispersion is activated is written as:Here, The force of gravity force is represented by:The external forces ag force . The theThermophoretic force makes particles move from higher to lower temperature regions, and stabilize in the corner of the computational domain.The particle temperature can be determined from the heat balance Equation :(29)mpCpThe convective heat transfer at the surface of the particle is expressed as:The heat transfer coefficient could be written using the Nusselt number:The thermophysical properties of silicon powders, reported in ,53, are ® Core (TM) i7-HP computer, with 1.99 GHz and 16 GB of installed memory (RAM), running a Windows 10 with 64 bit operating system. Simultaneous solutions of time-dependent coupled equations took 15 h 10 min.The turbulent thermal plasma jet transporting silicon nanopowder is simulated, using a computational fluid dynamic module and particle tracing module in three dimensions, via COMSOL Multiphysics 5.4 . The Gal2 (40/10 lpm)). The calculated temperatures are in good agreement with associated experimental measurements, which allows the validation of our model.In Following the plasma jet’s exit from the torch, the temperature profile shows a downward trend. The first rapid decline occurs due to the expanding process of plasma jet, no longer constrained by the torch nozzle. The second significant decline is associated with the shilling of the cold entrainment gas and cold powder carrier gas into the plasma jet central region.Numerical results of a turbulent plasma argon jet emerging in an argon atmosphere with particle injection are presented here. As a precursor material, silicon particles of 20 µm average diameter are freed simultaneously with argon gas in the plasma flame. Carrier gas and plasma mass flow ratio are assumed as 0.06 and 0.2 for an injection gas speed of 10 or 30 m/s, respectively. At first, the variation of calculated plasma jet temperature as a function of input power are presented in The swirl flow effect on velocity and temperature plasma field is represented in in = 10 m/s and vin = 30 m/s; see Plasma field velocity and temperature under different injection velocities are illustrated  injected in the plasma jet at different instants is studied , and jet characteristics.The impact of the injected carrier gas on the particle velocity and temperature distribution travelling along with the plasma jet at speeds of 15, 20, and 30 m/s are studied see . We demoThis result could be assigned to particle acceleration with the axial direction if the small effect of the external forces eglected :(34)dupdSince p = 10,000, vin = 20 m/s) with variable diameters  is shown in The path of silicon particles at the same instant . For a higher number of particles, total momentum and energy exchanges increase, and the injected particles assume deeper penetration during their motion in the gas channel  are illustrated in Here, the trajectories of the sprayed particles at different intensities of swirl flow (uThe effect of an injection of Si particles on the non-transferred turbulent argon plasma jet is investigated, using a three-dimensional numerical model. Plasma thermo-fluid dynamics, precursor-paths, particle growth process, and thermal history are described using a simulation model. The effect of different precursor flow rates, diameters, and the numbers of particles are examined, to determine the optimal growth conditions. The swirling flow reduces the localized vortex, and decreases transferred energy and momentum in the plasma jet. Consequently, particles deviate away from the plasma jet’s centerline. Temperature and velocity flow fields are strongly deformed at elevated carrier gas flow rates. An injection carrier gas velocity of 10 m/s is insufficient for the particles to be completely melted, while with an injected carrier gas velocity greater than 20 m/s, particles overshoot the central zone, and reach the outer slower region of the jet. The particle’s temperature and axial velocity vary drastically along the jet’s axis with the different sizes and numbers of the particles. At smaller diameters, particles attain higher terminal velocities, and accelerate through a longer flight distance. However, particles larger than 50 µm diameter remain below their melting point when sprayed. This is because, in this case, a larger amount of energy is needed to melt the particle. Furthermore, the initial plasma field distribution at the upstream flame influences the magnitude of the particle’s temperature and velocity distribution. A high-velocity plasma jet provides a short residence time, allowing the particle to be pulled into the hot core of the flame, and increases the heat transfer efficiency from the flame to the particle surface. Raising the plasma gas temperature increases the particle temperature strongly, despite the reduced residence time, due to high velocities. A plasma temperature of 8000 K is insufficient to melt the silicon particles.Therefore, thermal plasma plays a major role in controlling high-enthalpy flow, rapid quenching, and the ability to synthesize high-quality nanoparticles. Adjusting the optimal parameters, such as injection velocity of carrier gas about 20 m/s, particles with diameter 20–40 µm, thermal plasma jet issuing from non-transferred plasma torch with a relative velocity of 150 m/s, and relative temperature greater than 8000 K, as well as adding swirling flow to control the plasma jet fields deflection, suppresses locally strong deformation.


Bacillus thuringiensis svar. israelensis (Bti) larvicides are effective in controlling Aedes aegypti; however, the effects of long-term exposure need to be properly evaluated. We established an Ae. aegypti strain that has been treated with Bti for 30 generations (RecBti) and is still susceptible to Bti, but females exhibited increased susceptibility to Zika virus (ZIKV). This study compared the RecBti strain to a reference strain regarding: first, the relative transcription of selected immune genes in ZIKV-challenged females (F30) with increased susceptibility detected in a previous study; then, the whole transcriptomic profile using unchallenged females (F35). Among the genes compared by RT-qPCR in the ZIKV-infected and uninfected females from RecBti (F30) and the reference strain, hop, domeless, relish 1, defensin A, cecropin D, and gambicin showed a trend of repression in RecBti infected females. The transcriptome of RecBti (F35) unchallenged females, compared with a reference strain by RNA-seq, showed a similar profile and only 59 differentially expressed genes were found among 9202 genes analyzed. Our dataset showed that the long-term Bti exposure of the RecBti strain was associated with an alteration of the expression of genes potentially involved in the response to ZIKV infection in challenged females, which is an important feature found under this condition. Aedes aegypti can transmit several arboviruses, such as dengue, chikungunya, and Zika, which have a significant impact on global public health [Ae. aegypti populations is considered a global challenge [Bacillus thuringiensis svar. israelensis (Bti) are one of the most effective tools to fight mosquito and black fly, owing to its high activity and selective action, allied with its environmental safety [c health . The conhallenge ,3, and ihallenge . The micl safety ,6,7,8,9.l safety ,11,12,13l safety ,15. Aedes populations permanently established in urban areas [Ae. aegypti populations, we established a laboratory strain named RecBti. This strain has been exposed to Bti for 35 generations in order to mirror the conditions of chronic exposure to this larvicide [1 to F30 were susceptible to Bti and its toxins in comparison with a reference strain [30) individuals, did not display alterations compared with a reference strain, but females subjected to artificial oral infection with Zika virus (ZIKV) showed increased susceptibility [The multiple protoxins found within Bti crystals and their unique mode of action on the larval midgut are the major factors that counteract the selection of resistance, and no consistent report of resistance to such crystals has been documented to date ,17,18,19an areas . To invee strain ,24. Anottibility . Higher tibility .Together, these findings showed that the RecBti individuals displayed similar susceptibility to Bti and other biological parameters described above, compared with the reference strain, except for the increased susceptibility of females to ZIKV . We hypo30, which showed an increased ZIKV susceptibility in a previous study [35 larvae to Bti was also assessed by comparing it with the Rockefeller reference strain, as part of the monitoring of this colony during its long-term exposure to Bti [35 females unchallenged with ZIKV was compared with the RecL strain.This investigation is part of a longitudinal study of the RecBti strain, which has been chronically exposed to Bti. Previous findings ,25 and ae to Bti . Finally30 or F35 generations were investigated, as specified in each section. RecL was the reference used, except for the evaluation of Bti susceptibility, which was based on the international reference adopted for this specific purpose, which is the Rockefeller strain. RecL is a local reference strain that has been maintained in the insectary without exposure to any control agent since 1996 [®). Adults were fed with a sucrose solution (10%) ad libitum and females were also artificially fed with defibrinated rabbit blood once per week.RecBti and the two reference strains used, RecL and Rockefeller, were maintained in the insectarium of Aggeu Magalhães Institute at the Oswaldo Cruz Foundation (IAM-FIOCRUZ). RecBti was established in 2011 from the same geographic area as RecL ; at every generation, around 9500 third-instar larvae were treated with a Bti-based larvicide that caused around 74% mortality during the pre-imaginal phase. The adults that survived the Bti treatment during their larval phase were used as the next parental generation. The procedures of Bti exposure were fully described in Carvalho et al. . In thisnce 1996 . The Roc30 RecBti females had a significantly increased susceptibility to ZIKV, compared with the reference RecL strain [30 females were not exposed to Bti during their larval phase, and the last treatment was performed against F29 larvae. The transcription profile of seven genes , following the manufacturer’s instructions and specific primers for each gene  and the relative quantification was performed using Applied BiosystemsTM Analysis software, Relative Quantification Analysis Module v.3.3 [−ΔCtZIKV/2−ΔCBlood. Afterward, a comparison between strains fed with ZIKV-infected and uninfected blood was performed using 2−ΔCtRecBti/2−ΔCRecL. Then, the means and standard errors from two biological replicates of each strain (RecBti and RecL) and condition  were compared. For the validation of the DEGs, the means and standard errors from three biological replicates of each strain were analyzed. These analyses were carried out using Student’s t-tests in GraphPad Prism v.5.0.0 for Windows , considering a p-value < 0.05 to be statistically significant.Quantitative reverse transcription polymerase chain reaction (RT-qPCR) assays were performed for two purposes: first, to evaluate the transcription of the seven selected genes related to the immune response described in ach gene . The ribectively . The samle v.3.3 ,34. For 35 larvae to Bti, Cry11Aa, and Cry4Ba protoxins was investigated, prior to the RNA-seq of F35 females. For that purpose, bioassays were performed to determine the Lethal Concentrations (LCs) of these compounds for 50% (LC50) and 90% (LC90) after 24 h, compared with Rockefeller reference larvae [The susceptibility of the respective Fe larvae . Briefly35) and RecL females, tests were run with RNA samples from three pools of twenty nulliparous 7- to 10-day-old females from each strain. The females from both strains were not blood fed, and the RecBti females were not exposed to Bti during their larval phase. Total RNA samples of whole females from both strains were obtained using the RNeasy Mini Kit™  following the manufacturer’s instructions. RNA purity and concentration were determined using NanoDrop 2000TM and Qubit 2.0 (Thermo Fisher Scientific). RNA integrity was verified by visualization of samples separated by electrophoresis in 2% agarose gel. Paired-end libraries were prepared from total RNA using a TruSeq™ Stranded mRNA Library Prep Kit , following standard procedures, and sequenced using a MiSeq Reagent Kit V3™ (Illumina) for 150 cycles on an Illumina MiSeq Sequencer™ (Illumina) at IAM-FIOCRUZ.To compare the constitutive transcriptomic profile between the RecBti , and we found that, on average, all the bases had Phred scores higher than 30; thus, the removal of low-quality reads was not needed. Each library was then mapped against the genome assembly of the Aedes aegypti strain (Liverpool strain AaegL5) available in the VectorBase database accessed on 12 November 2019 (http://www.vectorbase.org). For the mapping step, the STAR aligner v.2.5.3 [p-values, in order to control the false discovery rate (FDR). To perform an exploratory analysis with the data, first, the expression data were used as input for the plotMDS function of the EdgeR package. EdgeR was used to perform differential expression analysis, considering only genes with one count per million (CPM) for at least all replicates of a group of samples (reference or RecBti strain). Genes with absolute values of log2 fold change (LFC) ≥1, and with FDR-corrected p-values ≤ 0.05, were considered to be DEGs. We selected the DEGs and plotted a scaled heatmap using log2 CPM with the heatmap R package. We created an MA plot that highlighted the DEGs. To further explore and refine the annotation of several DEGs tagged as hypothetical proteins in the Ae. aegypti genome, three additional annotation steps were performed using the Blastx tool with default parameters. The mRNA nucleotide sequences from all DEGs were used as queries against the most recent version of the following databases: (i) UniProtKB curated database; (ii) non-redundant database from NCBI, retaining only the top hit for annotation; and (iii) non-redundant database from NCBI using Blast2GO [Ae. aegypti using AnnotationHub version 3.2.2 [The quality of the sequenced reads was evaluated by applying the FastQC tool version 0.11.5  and domeless (AAEL012471) genes from the JAK-STAT pathway, which can stimulate the antiviral response, were investigated. In the comparison of infected and uninfected females analyzed within strains, both genes were significantly downregulated in RecBti, while they were upregulated in RecL (domeless and hop were significantly downregulated in RecBti (relish 1 (AAEL007696) and cactus (AAEL000709) were also evaluated. The gene relish 1, a target that stimulates viral defense, was significantly downregulated after viral infection in both strains, compared with their respective non-infected counterparts. When the status of infected females from the strains was compared, the expression was reduced in the infected RecBti females, although this was not statistically significant  and cecropin G (AAEL015515), no expression was detected in the RecBti-infected individuals  was the only AMP among the peptides tested that displayed detectable expression levels for both strains and conditions. Within strains, gambicin was repressed in the infected RecBti females, as well as between infected strains , which can be activated by the IMD or toll pathways after ZIKV infection in mosquitoes ,40, wereividuals . Therefo strains . This suividuals . While confirmed . The gen strains . The res35 females unchallenged with ZIKV described in 35 larvae to Bti and its protoxins. This evaluation was part of the continuous monitoring of this phenotype during the maintenance of this strain under Bti exposure. The RecBti F35 larvae were susceptible to Bti according to the LC50 and LC90 values, which displayed only discrete variations, compared with the Rockefeller reference larvae  and confirmed the susceptible status of the larvae to these compounds.Prior to the assessment of the transcriptome using Fe larvae . The tox0 larvae . LikewisAe. aegypti strain to Bti for 34 generations was related to a distinct gene expression profile in RecBti females (F35) that were not challenged with ZIKV. These females were compared with females from the reference strain (RecL), obtained under the same conditions, using a next-generation sequencing approach. For this purpose, F35 larvae were not treated with Bti and the respective females were not fed on blood. RNA samples obtained using pools of whole females from each strain, in triplicate, were subjected to transcriptome shotgun sequencing (RNA-seq). Analysis of the sequenced RNA-seq libraries showed a total of 10,103,560 and 13,058,581 reads obtained for the RecBti and RecL strains, respectively, and 87% and 85% of these genes were specifically mapped against the Ae. aegypti reference genome , adult cuticle (AAEL014978), glutamate decarboxylase (AAEL001902), timeless protein (AAEL019461), and alpha-amylase (AAEL013421), exhibited a profile of reduced expression; however, these were not statistically significant  toxin (RR50 > 5000) from Lysinibacillus sphaericus larvicides. The Bin-resistant larvae, also unchallenged with this toxin, showed a remarkable constitutive profile of differential expression characterized by 1300 DEGs, including the repression of several genes with LFC values greater than 3, including the gene coding the receptor of the Bin toxin, which is responsible for resistance [In another step of our study, the transcriptome of RecBti females was assessed to offer a broader view of the expression status of genes that could be differentially expressed by RecBti Fntil F30 ,25 and ontil F30 ,19. The d to Bti ,60,61, oi toxins ,63, in ge traits ,25. A cosistance . Bacillus thuringiensis [Cx. quinquefasciatus populations from Florida, with different susceptibility to West Nile virus (WNV), showed that the most susceptible individuals had greater expression of vitellogenin and bigger ovary size after blood feeding. Such a finding suggests a correlation between ovary development and competence to WNV [Ae. aegypti strain to Bti did not impact its susceptibility to this larvicide.Although the status of susceptibility to Bti was unaltered for the RecBti strain, which is consistent with the transcriptomic profile found for genes involved in the action of Bti, the increased susceptibility to ZIKV was associated with only two repressed DEGs related to arbovirus response; namely, “poor Imd response” and “leucine-rich immune protein,” and the repressed status of the former was not validated by RT-qPCR. On the contrary, the upregulation of vitellogenin-related genes in RecBti females should be noted, as vitellogenin can play an important role not only in reproduction, but also in immunity, including antibacterial action against ngiensis ,66,67,68e to WNV . It shoue to WNV ,72,73,74Ae. aegypti to the microbial larvicide Bti did not evolve to an alteration of the susceptibility to this larvicide, but it was associated with a repressed transcription status of immune genes in ZIKV-challenged females, which is consistent with their increased susceptibility to arbovirus. The whole transcriptome of RecBti unchallenged females did not reveal a remarkable set of differentially expressed genes, including most genes related to immune response, under the constitutive condition. Therefore, our dataset indicates the potential influence of long-term exposure to microbial larvicides in the modulation of transcripts potentially involved in the response to ZIKV infection in challenged females, and the mechanisms involved require further investigation, considering the importance of this feature found to be associated with this condition. Long-term exposure of the RecBti strain


The meta-analyses were performed using the generic inverse variance in R software with a random-effects model. Subgroup analysis was used to determine if the lung function was influenced by a potential study effect or by the participants’ characteristics. A total of 5562 participants from 24 studies were included. No significant difference was found between firefighters’ predicted FEV1 from wildland, 97.64% , and urban fires, 99.71% . Similar results were found for the predicted FVC. Nevertheless, the mean values of firefighters’ predicted lung function varied significantly among studies, suggesting many confounders, such as trials’ design, statistical methods, methodologies applied, firefighters’ daily exposure and career length, hindering an appropriate comparison between the studies.Firefighters are exposed to a range of harmful substances during firefighting. Exposure to fire smoke has been associated with a decrease in their lung function. However, the cause–effect relationship between those two factors is not yet demonstrated. This meta-analysis aimed to evaluate the potential associations between firefighters’ occupational exposure and their lung function deterioration. Studies were identified from PubMed, Web of Science, Scopus and Science Direct databases (August 1990–March 2021). The studies were included when reporting the lung function values of Forced Expiratory Volume in 1 s (FEV Exposure to fire smoke represents a severe health risk and is a growing concern for occupational and community exposures . Fire smIn general, individuals exposed to fire smoke can undergo different health effects, both physical and mental, with varying severity levels. Some of the hazards and concerns for public health are airway and eye hypersensitivity, changes in vascular, pulmonary and cardiopulmonary function, different types of cancer and, in more severe cases, death ,7,8,9,10The most vulnerable groups of the population include children, the elderly, pregnant women, individuals diagnosed with chronic cardiopulmonary diseases and occupational groups, especially emergency workers such as firefighters (including volunteers), police, rescue workers and health workers ,11. Fire1 and FVC [In an attempt to understand the impacts of fire smoke exposure on firefighters’ health, some studies evidenced that fire smoke may cause pulmonary embolism, pneumonia, bronchitis, chronic obstructive pulmonary disease (COPD) and asthma, as well as their exacerbations ,13,14,15 and FVC . However and FVC ,18. On t and FVC ,20,21. T and FVC ,23.The majority of those studies in the literature described urban fires, mostly related to the collapse of the World Trade Centre (WTC) on 11 September 2001 (9/11) ,25,26, wAs far as known, only systematic reviews have been published in this area. Thus, this study aimed to perform a meta-analysis to understand and quantify the impact of fire exposure on firefighters’ pulmonary health.Four databases, namely, PubMed, Web of Science, Scopus and Science Direct, were accessed in March 2021, using the same search terms: “firefighter”, “health effects”, “spirometry”, “asthma”, “occupational exposure”, “obstructive airway”, “lung”, “FEV”, “forest”, “wildland”, “chronic obstructive pulmonary disease” and “meta-analysis”. References cited in individual or review articles were systematically analysed through a manual search and included in this meta-analysis. No language limitation was applied. Duplicates were removed. The authors were not contacted for further information. This meta-analysis followed the PRISMA  guidelines .1 and FVC values, were included. The primary search results were reviewed, and some of the articles were eliminated after reviewing their title and abstract.Only cohort or case–control studies, which specifically evaluated the lung function and reported FEV1 or FVC; (v) overlapping study population; and (vi) published in books/book chapters, reviews, textbooks and reports.The applied criteria for exclusion of a study were: (i) not related to lung function; (ii) not related to firefighter workers; (iii) not related to fire exposure; (iv) not reporting FEVThe studies were independently researched and screened by two authors. Agreement between them was reached after a consensual discussion. For studies considering the same or overlapping populations, we selected the one with a larger population and more comprehensive information.1 (%) and FVC (%) were collected as indicators of lung function. When FEV1 and FVC were expressed in litres, the corresponding predicted values were estimated using the Global Lung Function calculator, which is based on the age, gender, height and ethnicity of the study participants [After applying the exclusion criteria, the studies were analysed, and data were extracted with the following information: author’s surname, publication year, study design, study location, sample size, participants’ mean age, fire type and main objective. Predicted FEVicipants .The missing standard deviations (SD) data were calculated from trial statistics, namely, confidence intervals and standard errors of the mean .2), Tau squared (τ2) and the standard chi-squared test (χ2). I2 is the proportion of the dispersion of the results observed in the studies included in a meta-analysis that is real, rather than specious. The I2 index can be considered as the percentage of the total variability in a set of effect sizes due to true heterogeneity (variability between studies). τ2 is the variance of the effect size parameters between the populations of studies, reflecting the variance of the true effect sizes. χ2 tests the statistical hypothesis that the true treatment effects (the parameters of effect size) are the same in all the primary studies included in a meta-analysis [2 values > 75%, moderate for values between 50% and 75% and low for values < 25%. The level of statistical significance was set at 0.05.Meta-analyses of the lung function values were carried out using the generic inverse variance in R software version 3.6.1 , with the meta package . Data ananalysis . HeterogSubgroup analyses were used to determine if the lung function was influenced by a potential effect of the study or by participants’ characteristics, such as publication year , study location , participants’ age , smoking practices (non-smokers and smokers) or type of fire (wildland or others).For the subgroup analysis by smoking practices, the studies were divided according to the number of smoking participants included in each study, with “non-smokers” and “smokers” assigned to the studies wherein less than or equal to 10% of the participants were non-smokers and more than 10% of the participants were smokers, respectively. Hereafter, whenever these terms are mentioned, it is intended to be understood as smoking and non-smoking participants.The methodological quality and the risk of bias for each of the included studies were assessed by two independent authors based on the Study Quality Assessment Tools . This to1 values were not available or were not eligible. Finally, the 24 studies obtained were divided into wildland (9 studies) and others/urban fires (15 studies). The PRISMA flow diagram is provided in After removing duplicates and including the records identified through manual reference analysis, the literature search identified 4297 studies. After screening the titles and abstracts, 4205 studies were excluded according to the exclusion criteria. Regarding specifically the criterion of overlapping study populations, several studies related to the WTC fire were found. However, as the analyses were performed based on the same population of firefighters exposed to the pollution of the WTC fire, only one study (the most complete) was considered in this meta-analysis. After reading the full text of the 92 remaining studies, 68 more were excluded, since the FEV1 mean values reported in the studies included in this meta-analysis varied from 82.94% to 113.39%. The overall predicted FEV1 mean value obtained for the 24 studies was 99.23% (95% CI: 94.65–103.80%), with a heterogeneity of I2 = 100%  in the studies performed before 1996, to 103.34%  in the studies published after 2014. This evident increase over the years was also observed in the predicted FVC mean values , while the participants from Australia showed the higher mean value of lung function, 105.19% . There were no significant differences between the participants from Europe, 98.11% , and those from North America 99.18% . The results obtained for the predicted mean values of FVC with the participants from Australia were also the highest, 111.10% ; however, the FVC values of the study performed in Asia, 103.90% , were slightly higher than those registered both in Europe, 102.65% , and in North America, 101.60%   .1 in firefighters according to their age. The youngest individuals (groups 20–30 and 31–40 years old) showed similar predicted FEV1 mean values , while the oldest participants (older than 40 years) showed a higher predicted mean value of FEV1, 101.95% . The same behaviour was observed for the predicted mean values of FVC (s of FVC .2 = 98%) and 97.03% , respectively  and those fighting other/urban fires . Similarly, there was no difference in the predicted FVC mean values  and missing or unclear information about the participants’ selection (inclusion/exclusion criteria). Finally, only four studies defined control groups.Two articles were rated as low-quality, 14 studies as moderate-quality, and 8 studies as high-quality . Conside1 and FVC mean values. Moreover, no statistically significant difference was observed in the pooled predicted FEV1 mean value in firefighters . This hindered definitive conclusions about the impact of firefighting exposure on the lung function. Still, the subgroup analysis allowed relevant interpretations.The results from this meta-analysis were obtained from 24 studies conducted along 31 years, including studies with distinct designs, in various countries and considering different types of fires and firefighters of different ages and with different smoking practices. The results showed a large variability in the predicted FEV1 and FVC values) was observed, although not statistically significant. However, the comparison between newer and older studies is difficult, as the studies’ procedures may have changed with time, namely, the recruitment strategies, the equipment used for lung function tests (past vs. modern devices), different or newer methodologies, or different standards for lung function tests [1 and FVC), which have been reviewed or modified over the years. The Global Lung Function Initiative developed in 2012 (GLI-2012) reports the latest reference equations for spirometry, based on the pooled resources of various countries and on data obtained from more than 74,000 healthy non-smokers tests from all over the world [Regarding the publication year, an increase in firefighters’ lung function along the years . Schermer et al. (2010) and Slattery et al. (2017) also reported high FEV1 values and associated them with the “healthy worker effect”. This effect describes the reduction of morbidity or mortality associated with employment factors when occupational cohorts and the general population are compared [Although non-statistically-significant differences were observed between different studies’ location, low pooled predicted FEVcompared ,64. In acompared . On the compared ,62.The lung function did not significantly vary with the firefighters’ age. It is known that with age, individuals undergo anatomical and physiological changes , which are responsible for the reduction in their lung function . Scherme1 predicted mean values compared to those of non-smokers was observed, though not statistically significant. Jacquin et al.’s study (2011), which was the wildland study with the highest score in the quality rating/lowest risk of bias assessment (0.92), reported that firefighters are likely to develop respiratory impairments after fire smoke exposure; however, the authors did not observe any statistical differences between smokers and non-smokers.Smoking is a known significant confounder. A slight reduction in smoking firefighters’ FEV1 mean values of both wildland and urban firefighters. Although this meta-analysis included 5562 participants, only 693 were firefighters dedicated to wildland fires, which probably may have contributed to overestimating the results obtained. Otherwise, some studies from North America assessed the lung function of firefighters who participated in WTC rescue operations. The collapse of the WTC generated a high-intensity pollution discharge, including extremely high Particulate Matter (PM) concentrations, exposing the population to extremely hazardous physical and chemical pollutants [The pooled analysis was unable to demonstrate a significant difference in the predicted FEVllutants . After tllutants . On the Heterogeneity was always above 90%. Perhaps, other factors that were not taken into account by most of the authors in the studies reviewed may have affected the results of the individual studies. Those include the duration of the intervention, i.e., the period between the end of the exposure and the lung function assessment (spirometry), the type of fire, and issues directly related to firefighters, namely, their age, other types of daily exposure besides that of firefighting, the annual number of working days and the career length.Other factors that may have contributed to not having found significant differences in lung function reduction was the use of respiratory protective equipment and firefighter’s type of career. The protective apparatus attempts to minimize the respiratory hazards which firefighters are exposed to, filtering particulates from the surrounding air or providing breathable air when working in oxygen-deficient or toxic atmospheres . On the Therefore, the results of this meta-analysis should be interpreted with caution due to the potential heterogeneity between the trials.1 and FVC).There was significant heterogeneity in the studies’ aims, designs, statistical methods and methodologies applied, which can influence the quality of the results or lead to variations in the reported values of the lung function parameters , and vice-versa, may have introduced bias in this study. Nevertheless, this is the most used methodology allowing an appropriate comparison between studies.The use of the GLI-2012 equations to transform the FEVFurther, due to the differences observed in how the data were reported, particularly, data measured in different units due to the use of different methodologies or data published in graphical figures, impairing the data extraction, it was necessary to exclude some studies or to perform recalculations (whenever possible) through the equations. Thus, this fact may have contributed to reduce the accuracy of the results.1 rates (between 82.94 and 113.39%) hindered definitive conclusions. Further, the pooled analysis was unable to demonstrate a statistically significant difference in firefighters’ predicted FEV1 mean values. Several factors could have contributed to this result, such as the methodologies applied, the equipment used, the reference equations for spirometry , the differences in the recruitment strategies of firefighters according to their countries or the publication year, hampering an appropriate comparison between studies.The present study’s goal was to assess the potential associations between firefighters’ occupational exposure to fire smoke and its effect on the lung function. The large variability observed in the reported predicted FEVSeveral reasons for statistical heterogeneity were identified. Since the results of the studies included in this meta-analysis may not have taken into account the possible confounding factors, they should be interpreted with caution due to the potential heterogeneity between the trials.Although there are some limitations, this study’s conclusions are of upmost importance because they highlight the necessity for further studies to assess firefighters’ lung function, especially in those combating wildland fires. This will allow to understand the impacts of fire on firefighters’ health, which are still unknown, and develop strategies to protect them. Moreover, further development could also include an analysis of the composition of the pollutants in each study, enabling to understand if the fire smoke composition influences firefighters’ lung function reduction.