Mitochondrial bioenergetics and oxidative balance in in vitro arbovirus infection models: a systematic review

Introduction: Viral infections affect oxidative metabolism and may have repercussions on mitochondrial alterations, compromising cellular homeostasis. Objectives: To assess mitochondrial bioenergetics and oxidative balance in in vitro arbovirus infection models. Methods: The review was written in accordance with the PRISMA and submitted to the Open Science FrameWork platform with DOI 10.17605 / OSF.IO / 8ZFSW. Were used the Descriptors/MeSH (Arbovirus, Arboviruses, Arbovirus infections, Mitochondria, Oxidative stress and Reactive oxygen species) was carried out on the platforms: PubMed, SCOPUS, COCHRANE, Lilacs and Web of Science. The quality analysis of the studies was performed using the ARRIVE tool adapted to the CONSORT, followed by the KAPPA concordance test, were used 24 articles. Results: Results show morphological alterations in the mitochondria, such as swelling, fragmentation, and the appearance of membranes. Mitochondrial stretching was more intense in regions close to the convoluted zones, associated with changes in the genes of mitochondrial dynamics. Changes in oxidative stress biomarkers, antioxidant enzymes and ROS production were evident in most articles, except for those that used cells of immunological origin. Conclusion: Changes in mitochondrial bioenergetic can assist the virus in the replication process, however, these changes can result causing damage cell and of oxidative stress.

Mitochondria have several functions such as the detection of intracellular homeostasis disorders and regulation and transduction of signaling responses, especially during stress conditions (Tait et al.,2012). These organelles are responsible for the production of ATP, through the oxidative phosphorylation process, which occurs in the internal mitochondrial membrane through the respiratory chain, using oxygen as a substrate for consumption (Sheeran et al., 2017). During the transport of electrons through mitochondria, the formation of reactive oxygen species (ROS) also occurs, which in excess, when the action of antioxidant systems overlaps, can cause tissue damage and cell apoptosis (Pizzino et al., 2017). To combat excessive ROS production, the cell has antioxidant defense mechanisms (enzymatic and non-enzymatic). The enzyme system is composed mainly of the enzymes superoxide dismutase (SOD), catalase, and glutathione peroxidase (GPX), which have the function of reducing the levels of ROS, preventing oxidative stress (Birben et al., 2012).
In arbovirus infections, these mitochondrial functions can be affected, as shown in a study carried out with the Venezuelan equine encephalitis virus in which structural changes, excess ROS production, decreased mitochondrial membrane electrical potential, and induction of fission of the organelle were observed (Keck et al., 2017). In addition, other studies have already shown that viruses in the family Flaviviridae, such as dengue, (Olagnier et al., 2014) and West Nile virus (Gullberg et al., 2015) induced increased ROS production and oxidative stress in infected cells. In addition to these, another study carried out with CHIKV, a virus of the Togaviridae family, showed that the virus induces a decrease in the mitochondrial membrane potential and an increase in MDA levels. Knowing this, the current review proposes to evaluate mitochondrial bioenergetics and oxidative balance in in vitro arbovirus infection models.

Search strategy
The research is a systematic review. The search was carried out without restriction of publication period and language. This systematic review was elaborated according to the items of the preferential reports for systematic analysis and meta-analysis (PRISMA) (Moher et al., 2009) found in table 4.
The search for articles was carried out in the following platforms: Medline / PubMed (Online System of Search and Analysis of Medical Literature), SCOPUS, COCHRANE, Lilacs (Health Sciences of Latin America and the Caribbean), and Web of Science. The following descriptors and MeSH were used in the platforms: Arbovirus, Arboviruses, Arbovirus infections, Mitochondria, Oxidative stress and Reactive oxygen species. The following crossings were performed: "Arbovirus Infections AND Mitochondria", "Arboviruses AND Mitochondria", "Arbovirus AND Mitochondria", "Arbovirus AND Oxidative Stress", "Arboviruses AND Oxidative Stress", "Arbovirus Infections AND Oxidative Stress", "Oxygen-reactive Species AND Arbovirus", "Oxygen-reactive Species AND Arboviruses", "Oxygen-reactive Species AND Arbovirus Infections". The references cited by the articles were also reviewed to identify any possible additional publications. This review was submitted to the Open Science Framework with identification DOI 10.17605 / OSF.IO / 8ZFSW. Research, Society and Development, v. 11, n. 16, e266111637749, 2022 (CC BY 4.0)

3-4
Objectives 4 Provide an explicit statement of questions being addressed with reference to participants, interventions, comparisons, outcomes, and study design (PICOS).

4-5 METHODS
Protocol and registration 5 Indicate if a review protocol exists, if and where it can be accessed (e.g., Web address), and, if available, provide registration information including registration number.

5
Eligibility criteria 6 Specify study characteristics (e.g., PICOS, length of follow-up) and report characteristics (e.g., years considered, language, publication status) used as criteria for eligibility, giving rationale.

5-6
Information sources 7 Describe all information sources (e.g., databases with dates of coverage, contact with study authors to identify additional studies) in the search and date last searched.

5
Search 8 Present full electronic search strategy for at least one database, including any limits used, such that it could be repeated. 5 Study selection 9 State the process for selecting studies (i.e., screening, eligibility, included in systematic review, and, if applicable, included in the meta-analysis).

5-6
Data collection process 10 Describe method of data extraction from reports (e.g., piloted forms, independently, in duplicate) and any processes for obtaining and confirming data from investigators.

6
Data items 11 List and define all variables for which data were sought (e.g., PICOS, funding sources) and any assumptions and simplifications made. *

Risk of bias in individual studies
12 Describe methods used for assessing risk of bias of individual studies (including specification of whether this was done at the study or outcome level), and how this information is to be used in any data synthesis.

6
Summary measures 13 State the principal summary measures (e.g., risk ratio, difference in means). 6 Synthesis of results 14 Describe the methods of handling data and combining results of studies, if done, including measures of consistency (e.g., I 2 ) for each meta-analysis. * Source: Authors.

Eligibility criteria and exclusion criteria
After removal of duplicates, the full articles were analyzed when they met the following criteria: 1) Type of study, in vitro studies that did not carry out pharmacological intervention; 2) Studies that performed some analysis on mitochondrial bioenergetics and / or oxidative balance; 3) Studies that used an arbovirus for infection in cells; 4) Original studies.
Thus, the following were excluded from this research: Review articles, case reports, papers that did not address mitochondrial bioenergetics or oxidative balance, papers without comparison with control, and any other study that did not meet the eligibility criteria.

Methods for data extraction
Two reviewers independently extracted data from each included article. A pre-piloted and standardized form was used to extract data from the included studies to assess the quality and synthesis of evidence. The primary outcomes of the study were: oxidative stress biomarkers, mitochondrial morphology, activity of antioxidant systems, and production of ROS. Secondary outcomes were: type of virus, concentration of virus for infection, viral incubation period, and relationship of regression of infection over time.

Quality assessment of included studies
In the absence of a consensus standard for evaluating the quality of in vitro studies, we used the modified ARRIVE tool, combined with the CONSORT guidelines (Consolidated standards for evaluation reports) for in vitro experiments, based on previous studies. The checklist contains 12 items to evaluate. For each item, a judgment related to the article was assigned, taking into account a pre-specified question. Next, Kappa statistics were used to assess interobserver agreement for items at risk of bias, using the software Statistical Package for the Social Sciences -SPSS version 20 for Windows (IBM SPSS Software, Armonk, NY, EUA). The results were synthesized in the form of summary tables and a descriptive summary to enable their comparison.

Results
After searching the electronic databases, 1741 articles were found. After the evaluation of duplicates, 1206 articles were excluded and 535 were submitted to reading of the title and abstract. In total, 28 articles were read in full because they fit the eligibility criteria, of which 4 were excluded due to the exclusion criteria. All results were published in English. The details of this search are shown in the Figure 1.

Evaluation of the quality of studies
The results of the Kappa tests revealed an almost perfect agreement, Kappa = 0.82 for the items evaluated by the tool (table 1) by described Viera, Garrett, 2005. The biggest disagreements were found in the objective items, sample discrimination, and reporting of procedures to reduce bias (Banerjee et al., 2018;Cherupanakkal et al., 2018;Cavalheiro Mariana et al., 2016;Camini et al., 2017;Dhanwani et al., 2012;Fernandes-siqueira et al., 2018;Keck et al., 2017;Keck et al., 2018;Lai et al., 2018;Moreno-altamirano et al., 2015;Mukherjee et al., 2013;Narayanan et al., 2011;Narayanan et al., 2014;Olagnier et al., 2014;Silva da Costa et al., 2012;Tung et al., 2010;Valero et al 2013;Verma et al., 2016;Yu et al., 2015). Regarding the objectives, some articles did not make their objective clear, with a tendency when writing the articles to observe the objectives implicit in the main results mentioned in the introduction. Regarding the discrimination of the samples, a failure was identified in the articles to detail the concentration of cells for the analyses, as well as the MOI (multiplicity of infection) of viruses that would be used for infection, which can represent a risk of bias when the analyses are associated with the evolution of the infection over a period.
In addition, 13 of the 24 articles did not report any procedure to minimize possible bias in the study. Most of the articles presented a clear and adequate description of the results (n=17) and discussion (n=23) for the proposed study, indicating a good written 1) Identification: potential studies were collected from databases; 2) Screening: screening of the studies was carried out based on the reading of the titles and abstracts, according to the PICO scale; 3) Eligibility: studies were accepted according to the eligibility criteria for complete reading of the articles; 4) Articles selected after complete reading for data extraction and qualitative analysis. and argumentative relationship between the results and discussion, as well as clearly stating the values (mean, error, and standard deviation) of the results obtained.

Cell characteristics and virus titration used in the studies
The cells used in the studies, for the most part, were of human origin. The results show that 79% of the studies with in vitro virus infection used cells of human origin and the other 21% used neuronal cells from mice and / or rats. The diversity of cells of human origin was associated with the type of virus and its preference for the cell type. The majority of the studies, especially those with Flavivirus, used blood cells for the experiments (dendritic cells, monocytes / macrophages, lymphomononuclear cells). Regarding the concentration of cells used for the experiments, the lowest concentration used was 103cel / mL, (Keck et al., 2017) but most articles used the concentration of cells 106 cel / mL (Keck et al., 2017

Arbovirus Infection and Mitochondrial Bioenergetics
The results of the studies show that arbovirus infections cause a series of mitochondrial changes that arise as a result of viral action or metabolic alterations in cells. To facilitate the understanding of the virus and mitochondria relationship, the results are described according to the family of the virus studied (Table 2).
In relation to the family Peribunyaviridae, infection by the La Cross virus, showed that in neurons there is an increase in the expression of the SARM1 binding protein and that this protein has a strong relationship with mitochondrial damage. The neurons infected by the virus showed mitochondria in a state of degeneration and increased production of EROS. The increase in the production of EROS was also identified in infections by viruses of the Togaviridae and Flaviviridae family.
With respect to the Flaviviridae family, in infected cells, the mitochondria in the host undergo changes in the consumption of energy substrate, for example, in DENV infections, mitochondria use palmitate and glucose as an energy substrate, whereas in non-infected cells glutamine is the most commonly used substrate. In addition, studies show that DENV infection increases the oxidation capacity of fatty acids. In another study (Chatel-Chaix et al., 2016) showed that in infected cells there is an increase in the mitochondrial respiration process, indicating greater availability of ATP for the cell, which contributes Regarding the Togaviridae family, studies were found with MAYV, VEEV, CHIKV, and SINV viruses. Regarding MAYV, studies show that infected cells demonstrate an increase in the production of ROS after 6 hpi and that this production signals the immune system to produce TNF-α, indicating that there is a relationship between the immune response and the production of EROS. With respect to SINV, in intact cells, there is no change in O2 consumption during 15hpi, but there is a reduction in the electron transport system (ETS); in permeabilized cells, however, there is a reduction, although not statistically significant, in the capacity of oxidative phosphorylation of cells when using substrates of complex I or II, of the electron transport chain. Thus, the capacity of the ETS dependent on CI and CII was not significantly affected after 15 h of infection by SINV, contrasting with the results with intact cells, where a significant decrease in maximum uncoupled respiration was observed at this time. However, the data point to a decrease in the activity of the mitochondrial membrane potential of cells infected by CHIKV and VEEV. Along with this change there is an increase in DRP1, indicating an increase in mitochondrial fission, in addition to changes in infected mitochondria such as swelling, membrane compression, and aggregation of damaged mitochondria (Cavalheiro Mariana et al., 2016;Dhanwani et al., 2012;Keck et al., 2017;Silva da Costa et al., 2012). To assess the relationship of SARM1 in neuronal apoptosis that is caused by LACV infections .
• SARM1 protein is associated with mitochondrial damage; • Infected cells showed an increase in the expression of SARM1 binding proteins with the mitochondria.

•
Neurons containing swollen and / or damaged mitochondria in cell bodies and axons from cultures infected with LACV; • Blocking the mitochondrial antiviral signaling protein (MAVS), showed an inhibition of SARM1 activity and EROS production.

•
There is no blockage of MAVS when there is an increase in the production of EROS; • SARM1 interacts with MAVS and induces oxidative stress, mitochondrial damage, and neuronal death. Fernande s-siqueira et al (2018) Flaviviridae.
Dengue vírus (DENV) / MOI of 1 Huh7 cells, a strain of human hepatocarcinoma / The cells were seeded in 60 cm plastic Petri dishes, at a density of 10 5 cel / mL, but in experiments such as oxygen consumption, concentrations of 10 6 cel / mL were used Using high-resolution respirometry to assess metabolic circuits that occur during DENV infection

Infection of cells by DENV inhibits mitochondrial breathing; •
Infected Huh7 cells effectively use palmitate as an oxidative substrate, since oxygen consumption in the presence of palmitate was higher than that observed in the control; • Glutamine was the best substrate used by control cells in all conditions, however, DENV infection strongly inhibited the use of this nutrient as an energy substrate for cells;

•
The presence of glucose in the control cells inhibited consumption, but these effects were attenuated in cells infected by DENV, confirming that the use of glucose by these cells increases the oxidation capacity of endogenous fatty acids.

•
Using images of live cells from cultures infected with DENV, it was seen that the infection induces mitochondrial elongation; • The mitochondria of cells infected with DENV showed increased respiration, which can lead to an increase in ATP production, when compared to uninfected cells; • Viral proteins, such as NS4b, were preferentially located in subcellular fractions of mitochondria;  To determine whether murine microglia is susceptible to VEEV infection and whether the infection results in mitochondrial dysfunction The infected group showed an increase in the expression of the Drp1 protein from 24 dpi in MOI of 5 to 10, while in MOI of 20 there was a drop in levels.

•
Infected cells showed a series of changes in structure, including partially swollen mitochondria, inner membrane compaction, and assembled mitochondria.

Oxidative balance and Arbovirus infection
Oxidative balance is extremely important for maintenance of cellular homeostasis. The results show several changes in the levels of oxidative stress biomarkers, and expression and/or antioxidant enzymes activity, in addition to changes in the cellular redox state. Changes were verified in the ROS production in more than 80% of the articles of this review. All the results described with respect to oxidative balance are shown in Table 3.

Antioxidant enzymes and oxidative stress biomarkers
In the Bunyaviridae family, the results show that in infections with Rift Valley Fever Virus (RVFV), SOD activity decreases in infected cells, and when the SOD is blocked, apoptosis levels increase about seven times (Narayanan et al., 2011).
These results are similar to those of Banerjee and Mukhopadhyay (2018)  however when related to cell apoptosis there was a significant difference between the infected group and the control group. In contrast, Valero et al (2013) showed that monocytes infected by DENV show increased MDA levels (Valero et al 2013). These discrepancies may be associated with the methodology used to infect the cells depending on the strain type. Regarding the Togaviridae family, Banerjee and Mukhopadhyay (2018) showed that in infections by the Chikungunya virus, lymphomonuclear cells of patients with or without polyarthralgia have high levels of MDA and high levels of the MDA/SOD ratio, indicating a possible situation of oxidative stress in these cells (Banerjee et al., 2018).

ROS and NO production
The ROS production was evaluated in most studies, thus, the ROS levels of all virus families in this review were quantified. Only one article did not show an increase in intracellular ROS levels; in that article the neutrophils infected by DENV with an MOI of 10 did not demonstrate an increase in the ROS production. In addition, the author concluded that the production and neutrophil activity are not associated with ROS (Moreno-altamirano et al., 2015).
In the Bunyaviridae family, there was an increase in the ROS production in liver cells infected by RVFV, associated with the presence of non-structural viral proteins and a decrease in SOD levels, where the author reports that there is downregulation between the decrease in SOD levels and increase in ROS (Narayanan et al., 2011). In the family Flaviviridae, the ROS increase was found in infections by DENV and JEV, in several cell types. In addition, astrocytes isolated from rats and infected by JEV showed a relationship between the increase in ROS and the expression of metalloproteinase-9 (mmp-9) (Cherupanakkal et al., 2018;Datan et al.,2016;Qi et al., 2015;Tung et al., 2010;Valero et al 2013;Yang et al., 2010). Studies with Flavivirus have shown that an increase in ROS occurs between 18 to 48 hpi. The Togaviridae and Rhabdoviridae families also demonstrated increases in ROS in infections by VEEV, CHIKV, SINV, and CHPV (Keck et al., 2018;Verma et al., 2016).
Regarding NO production, two studies carried out experiments to quantify NO production. In the first study performed with DENV, it was observed that in infections by the 4 dengue serotypes there is an increase in NO production, however the highest production was found in DENV-2. In addition, the NO increase was associated with an increase in MDA (Valero et al 2013). The other study was carried out with CHPV, in which Verma et al. (2016) showed that at 16hpi there is an increase in NO production associated with ROS production in BV-2 cells (Verma et al., 2016). In addition, when the neuronal apoptosis index was evaluated, an increase in the cell death of the infected group was evidenced, indicating that NO and ROS production may be associated with apoptosis of infected cells. • The infected cells showed a consistent decrease in expression of superoxide dismutase 1 (SOD1) from the initial moments (24 and 30 h); • There was an increase in the ROS production in infected cells related to the negative regulation of cytosolic SOD; • The increase in the production of superoxides was visible from 12dpi and 24dpi; • Control cells treated with TNF-α showed lower levels of SOD1 compared to untreated cells, in a 24-hour period; • After depletion of SOD1, the percentage of cells undergoing apoptosis was almost seven times higher than the control cells; • SOD1-depleted cells that were not infected by the virus had significantly less apoptosis (1.5 times greater than control cells; data not shown); • The results indicate that SOD1 depletion makes the cells more susceptible to apoptosis during MP12 infection; • Activation of the AMPK pathway was necessary to combat the damage caused by changes in the SOD1 concentration; • Cells infected with the ZH501 strain of RVHV were subjected to the same experiments and demonstrated similar results, but the MOI used was lower than that of the MP12 strain due to its pathogenicity. Narayanan et al (2014) Bunyaviridae Rift Valley Fever Virus (RVFV) / MOI of 3.5 or 10 Liver stellate cell line (CFSC-8B) / 1.10 6 cell/mL To investigate whether oxidative stress occurs and whether it leads to apoptotic responses in patients infected with RVFV • Quantification of fluorescence revealed an approximate 6-fold increase in superoxide levels in virus-infected cells over controls infected by simulation; • The presence of the viral protein NSs in the mitochondria of infected cells contributes to the early increase of ROS; • ROS increased levels correlated with the activation of nuclear factor-κB (NFκB), p65 and p53 responses, which together with the infection were also reflected as macromolecular rearrangements; • ROS act as critical contributors to liver cell apoptosis during RVFV To evaluate the mechanism of autophagy in DENV infection at an early stage and how this process is related to the endoplasmic reticulum (ER) and the protein kinase of the type R endoplasmic reticulum (PERK) • The inhibition of ER stress or ataxia mutated telangiectasia (ATM) signaling cancels the protection provided by dengue against other cellular stressors; • Direct inhibition of the ER stress response in infected cells decreases the renewal of the autophagosomes, reduces the ROS production, and limits the dengue virus reproduction; • Blocking ATM activation, which is an early response to infection, decreases the transcription of ER stress response proteins, but the ATM has a limited impact on the ROS production and viral titers; • The ROS production determines only late-onset autophagy in infected cells and is not necessary for dengue-induced protection against stressors; • Among the various autophagy-inducing pathways during infection, ER stress signaling is most important for viral replication and cell protection. Valero et al (2013) Flaviviridae Dengue virus (DENV-1, DENV-2 and DENV-4) / MOI of 1 Monocytes / 3.10 5 cell/mL To analyze the oxidative (nitric oxide-NO and malondialdehyde -MDA levels) and antioxidant (catalase-CAT, superoxide dismutase-SOD and reduced glutathione-GSH) responses of monocytes of newborns, young adults, and older adults during in vitro study of dengue virus infection • Lower values of SOD, MDA, GSH, and NO were found in neonates and older adults, whereas the highest values were observed in adults; • The increase in NO, MDA production, catalase and SOD activities, and the GSH content were induced by different viral serotypes and LPS in monocytes of neonatal individuals, adults, and older adults; • DENV-2 induced the highest NO production accompanied by high MDA production; • The antioxidant response was observed in all types of DENV; • The highest values of catalase activity were observed for DENV-1 and DENV-4 and SOD activity for DENV-1; • The highest GSH content was observed in DENV-4 infected monocytes in all groups analyzed; • The degree of oxidative and antioxidant response was influenced by the source of monocytes. Qi Y et al (2015) Flaviviridae Dengue virus (DENV-2) / MOI of 4 Primary isolates from human umbilical veins (HUVECs) / 10 6 cell/mL To evaluate the interferon 6 (IFN-6) role in infection by DENV-2 • The decrease in mitochondrial membrane potential (MMP) was noticeable from 24 hours to 48 hours after DENV2 infection; • Infected IFN 6 cells (+/+) showed no changes in MMP for long periods of time. On the other hand, IFN6 -/-cells appeared to be more sensitive to DENV-2. Tung et al (2010) Flaviviridae Japanese encephalitis virus (JEV) / Not described Rat brain astrocytes (RBA-1)/Not described To evaluate the molecular mechanisms underlying JEV-induced MMP-9 expression in RBA-1 cells • Treatment with JEV induced a significant increase in ROS levels, measurable in 5 minutes and sustained for more than 90 min, which may be associated with an increase in NADPH oxidase; • JEV-induced MMP-9 expression was mediated through ROS-dependent activation of p42 / p44 MAPK in RBA-1 cells. Cherupanakkal et al (2018) Flaviviridae Dengue virus (DENV) / Not described

Lymphomonuclear cells /Not described
To assess biomarkers of oxidative stress, apoptosis, and DNA damage in individuals infected by dengue virus • Increased levels of lipid peroxidation were found in the infected group when compared to controls with febrile illnesses and controls without febrile illnesses; • When MDA data were correlated with apoptosis levels, a positive correlation was found according to Spearman's correlation. Keck  • Nitric oxide (NO) was 1.82 times higher in infected patients who had persistent polyarthralgia (ChikWPP); • The ChikWopp group had higher levels of thiols than ChikWPP; • Carbonyl levels were higher in patients with polyarthralgia; • SOD levels were lower than the control; • The SOD level was 1.02 times lower, while the radical scavenging activity was 1.16 times lower in infected patients than in controls; • The MDA/SOD ratio was 1.57 times higher in the ChikWPP group; • There was a 2.42-fold increase in intracellular ROS production when compared to the control group Research, Society and Development, v. 11, n. 16, e266111637749, 2022 (CC BY 4.0)  • The ROS increase corresponded to a decrease in the mitochondrial membrane potential (MMP) in both cell lines; • At 24 hpi, astrocytes infected with TC-83 exhibited a 53.6% decrease of the MMP compared to uninfected control; • Correlating with the lower ROS level, the BV2 microglia suffered a 40% decrease in MMP at 24 hpi; • The ROS accumulation and the MMP decrease were not dependent on the MOI and, therefore, all subsequent experiments were conducted using the MOI of 2; • The anti-inflammatory BAY-82 decreased the total ROS levels by 16% at 6 hpi and 22% at 24 hpi when compared to the control infected by TC-83. Cmini FC et al (2017) Togaviridae

Mayaro virus (MAYV) / MOI of 5
Human hepatocarcinoma cells (HepG2) or Macrophages (J774) / 2.5.10 4 cell/mL or 1.10 6 cell/mL To investigate whether MAYV causes oxidative stress in infected cells and affects the host's antioxidant system • There was an increase in total SOD activity at all times in cells infected with MAYV; • There was an increase in the CAT activity at 15 hpi, in cells infected with MAYV; • The total glutathione content in cells infected with MAYV increased by approximately 4 and 2.5 times at 6 and 15 hpi; • Total glutathione levels remained normal in cells at 24 hpi; • There was a progressive decrease in the GSH/GSSG ratio in HepG2 cells after infection by MAYV, with 35% and 37% reductions in this ratio at 15 and 24 hours after infection, respectively; • Similar to observations in HepG2 cells, ROS production was increased in MAYV-infected J774 cells; • Regarding HepG2 cells, there was an increase in ROS generation at all times tested (1, 2, 4, 6, 15, and 24 h pi), while in J774 cells an increase was observed after longer periods of infection (6, 15, and 24 h pi); • J774 cells showed a decrease in total SOD activity and increase in MDA levels at 6, 15, and 24 hpi Dhanwani R et al (2012) Togaviridae Chikungunya virus (CHIKV) / MOI of 0.1 to 5 Human neuroblastoma (SH-SY5Y) / 1.10 6 cell/mL To evaluate the role of apoptosis and oxidative damage in SH-SY5Y cells infected with CHIKV • There was a decline in the GSH level over time; • Substances reactive to thiobarbituric acid (TBARS) production reached a maximum at 48 hpi. In addition, the oxidative stress level increased in parallel with the viral titer, at all times. To evaluate the pattern of microglial activation and dosage of proinflammatory cytokines in CHPV infection • The ROS and NO levels were evaluated at different incubation times and increased concomitantly with time; • It was found that the NO level was higher at 16 hpi; • The ROS level in the sample infected by CHPV was higher at 24 hpi, compared to those infected by simulation; • Regarding cell death, neuronal cells treated with the supernatant obtained from CHPV-infected microglia showed a greater number of TUNEL positive cells, compared to cells infected by simulation.

Discussion
Viral infections have already been associated with imbalances in the oxidative metabolism of infected people, however, to date, no review has been carried out to identify how arbovirus infections act on REDOX status and mitochondrial activity in in vitro models. Therefore, the current study aimed to evaluate how arbovirus infections affect oxidative balance and mitochondrial bioenergetics in in vitro study models. Thus, analyzing the effects that infections cause in in vitro models, can serve as a basis for understanding the pathological mechanisms caused by arboviruses and directing pharmacological or nonpharmacological treatments to act more effectively in minimizing cell damage caused by infections.

Cell response to arbovirus infection
In general, all articles indicate an increase in the production of reactive oxygen species because of infection. The increase in the production of ROS was also associated with an increase in proteins such as sterile Protein α and TIR containing protein 1 (SARM1) as well as which, changes in the energy substrate were also identified, as a result of the process of infection and viral replication. The change in the energy substrate associated with an increase in the breathing process, and an increase in the MMP, indicate greater availability of ATP for the virus, which may indicate a viral stimulus to obtain energy through changes in cellular homeostasis. This became more evident when it was identified that the morphofunctional changes in the mitochondria occurred close to the CM, an area where viral replication is intense (Keck et al., 2018;Mukherjee et al., 2013).
Regarding the changes in mitochondrial morphodynamics, of the 24 selected articles, only 4 performed analyses related to mitochondrial dynamics, in which, in the consequences, it was identified that arbovirus infections increase the expression of Drp1, indicating a process of fission of mitochondria. In addition, two of the studies showed that the expression of MFN1 and MFN2 decreased with viral infection. In relation to the total number of articles found in the research, few assess the mitochondrial dynamics in infections; of the 24 articles, only 8% show analyses on the mitochondrial dynamics (Chatel-Chaix et al., 2016;Yu et al., 2015).
The mitochondrial elongation was quite expressive in the mitochondria; however, this was not the only change in morphology found; since the articles show that there are partially swollen mitochondria, and inner membrane compaction and mitochondria agglomeration. Therefore, mitochondria respond to viral infections by increasing their respiratory activity and generating morphofunctional changes, however, it was not clear in the studies whether these responses decrease the infectious process or if, due to the increase in the availability of ATP, they facilitate the viral replication process, or even if there were harmful changes to the cell, especially because of the increase in ROS.
Therefore, the increase in total cell ROS was evaluated by some studies, followed by analyses of some biomarkers of oxidative stress. One study stated that there was no increase in the production of reactive species in neutrophils infected by DENV. This result implies that there is no relationship between the increase in ROS and infection of neutrophils by the dengue virus. However, other studies show a relationship between the increase in ROS as a signal for an immune response. The study by Qi Y et al (2015), showed that in infections by DENV, the increase in ROS is associated with the production of IFN-6 and that this affects the cell's MMP (Qi et al., 2015). In addition, another study by Narayanan et al (2011) showed that there is a negative relationship between ROS, SOD, and the increase in TNF-α (Narayanan et al., 2011). Thus, although ROS does not stimulate the production of neutrophils, it is associated with the activation of some mechanisms related to the immune system, indicating that it may be one of the triggers for the activation of the immune response against viral infection. Regarding biomarkers and enzymes, viral infection induces an increase in the activity of SOD and CAT enzymes. In the studies in which SOD levels were reduced, there was an increase in MDA associated with an increase in apoptosis of infected cells. In addition, the levels of GSH and the GSH / GSSG ratio were also reduced in viral infections. In short, the reduction in the activity of the antioxidant defense system is associated with an increase in the MDA biomarker, indicating a possible framework of oxidative stress during infection. In addition, MDA levels were also associated with increased NO in infected cells and increased apoptosis.

Study information
The availability of articles that evaluated the damage to mitochondria and the oxidative balance of infected cells is quite scarce when considering the diversity of pathogenic arboviruses that affect humans. Most studies are related to Flavivirus or Togavirus, such as Dengue and Chikungunya; this result may be associated with the importance that these viruses have for public health, principally in this century (WHO, 2014). However, some arboviruses that were also of great importance in this century were not the subject of in vitro studies and research that met the eligibility criteria of this work, for example, none of the studies available with the Zika virus were found in the search.
In the majority of the studies, the cell infection process is not very detailed and, in some articles, important parameters such as infection time, virus MOI, cell concentration, and culture medium details were not identified. These results can serve as a bias for comparing the work, where the evolution of the infectious process may be associated with the concentration of virus used.
In addition, another limitation of the studies was the use of the term "oxidative stress" in studies that evaluated, solely, the production of reactive species or a single biomarker of oxidative stress, therefore, few methodological analyses were performed to indicate that the cells really were experiencing oxidative stress.

Final Considerations
The results of this review indicate that in in vitro arbovirus infection models changes in the mitochondrial bioenergetics can help the virus in the replication process, however, cellular changes resulting from the exaggerated increase in reactive species cause an imbalance in the cell REDOX state which can lead to oxidative stress and cell death signaling.
Based on these results, research is suggested to understand how the impacts of ROS caused by arboviruses in in vivo models to understand systemic repercussions in the organism, as well as the role of EROS in the etiology of arboviruses.