Evaluation of long noncoding RNA (LncRNA) in pathogenesis of HELLP syndrome: diagnostic and future approach

Abstract HELLP syndrome is a disorder during pregnancy which is defined by elevation of liver enzymes, haemolysis, and low platelet count. This syndrome is a multifactorial one and both genetic and environmental components can have a crucial role in this syndrome’s pathogenesis. Long noncoding RNAs (lncRNAs), are defined as long non-protein coding molecules (more than 200 nucleotides), which are functional units in most cellular processes such as cell cycle, differentiation, metabolism and some diseases progression. As these markers discovered, there has been some evidence that they have an important role in the function of some organs, such as placenta; therefore, alteration and dysregulation of these RNAs can develop or alleviate HELLP disorder. Although the role of lncRNAs has been shown in HELLP syndrome, the process is still unclear. In this review, our purpose is to evaluate the association between molecular mechanisms of lncRNAs and HELLP syndrome pathogenicity to elicit some novel approaches for HELLP diagnosis and treatment.


Introduction
HELLP is the syndrome defined by elevation of liver enzymes, haemolysis, and low platelet count (Khalid et al. 2022).According to recent studies, this syndrome occurs in almost 0.2% of pregnancies (McCormick et al. 2022).Although the exact features of pathogenicity for this syndrome is still unknown, some symptoms have been detected such as maternal vascular malperfusion, which is formed by thrombosis of decidual arterioles that lead to placental infarction and villous hypoplasia (Perez Botero et al. 2021).Many studies revealed that this syndrome is a multifactorial one and both genetic and environmental components can have a crucial role in this syndrome's pathogenesis.In addition, it has been shown that using different diagnostic methods to evaluate tissues can help in their management (Molvarec et al. 2008, Khalid et al. 2022, Liu et al. 2022c).
Over the last decade, much research about RNAs showed that these elements are multifunctional and enable to regulate many cell processes.Recently, subcellular localisation of RNAs has highlighted as a common fact which affects many cellular procedures.Long noncoding RNAs (lncRNAs) are defined as long non-protein coding molecules (more than 200 nucleotides), which are functional units in most cellular processes.It is also important to evaluate lncRNAs' subcellular localisation to perceive their biology.According to some research, these factors play a vital role in cell cycle, differentiation, metabolism and also some diseases progression (Yan et al. 2020, Zhuo et al. 2020, Bridges et al. 2021).As these markers discovered, there has been some evidence that they have an important role in the function of some organs, such as placenta (McAninch et al. 2017).Regarding to some research done about the association of lncRNAs and some disorders such as HELLP, investigators detected that alteration and dysregulation of these RNAs can develop HELLP disorder (Abdollahpour et al. 2018, Zhang et al. 2020, Omertayeva et al. 2021).Indeed, the role of some lncRNAs in pregnancy-specific HELLP syndrome is mediating the splicing mechanism; however, this process is still unclear (Pisignano and Ladomery 2021).LINK-HELLP is a lncRNA within both nucleus and cytoplasm of first extra villous cytotrophoblast cells.Mutagenesis research about this lncRNA has indicated its important role in invasion and trophoblast proliferation.It is also mentioned that knocking down of LINK-HELLP lncRNA leads to gene expression reduction that may demonstrate the association of lncRNAs with transcriptional or posttranscriptional regulation by RNA splicing (McAninch et al. 2017).
Our purpose in this review is evaluating the molecular role of lncRNAs in HELLP pathogenicity to elicit some novel approaches for HELLP diagnosis and treatment.

IL-6
HELLP syndrome defined as a disorder which is affected by inflammatory factors.During pregnancy along with HELLP syndrome, the placenta has been detected as the primary source of Fas ligand (FasL) which has an impact on inflammation.According to recent studies, neutralising of FasL can lead to a decrease in inflammation in pregnant women with HELLP syndrome (Gibbens et al. 2020).Stimulation of Fas can induce the expression of some pro-inflammatory elements including matrix metalloproteinase (MMP)-9 and IL-8.It is also indicated that the Fas signalling is initiated from mediation of mitogen activated protein kinases (MAPKs) including extracellular-signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) which stimulate subsequent activation of nuclear factor kappa light chain enhancer of activated B cells (NF-jB) (Lee et al. 2011b).In some research, miR-204-5p was detected as the inhibitor of FasL.Therefore, this miRNA can regulate Fas/FasL pathway.Indeed, this miRNA exerts its role via IL-6/IL-6 receptor pathway regulation.IL-6R is the target of miR-204 and a regulatory loop is made between this miRNA and IL-6R/STAT3 signalling pathway.Therefore, miR-204 can repress the IL-6R/STAT3 pathway which causes inflammation repression (Li et al. 2018a, Li et al. 2022, Zhu et al. 2019).Even though the role of different kinds of lncRNAs in HELLP syndrome is still unclear, there exists some evidence showed the role of these markers in HELLP syndrome progression.In the study by Zhizhong et al. the role of lncRNA nuclear paraspeckle assembly transcript 1 (NEAT1) is demonstrated in modulating miR-204-5P expression level.This lncRNA induces Tumour Necrosis Factor alpha (TNFa) and subsequently activation of NF-jB signalling pathway by down-regulation of miR-204-5p (Xiao et al. 2021).According to this investigation, lncRNA NEAT1 may have a role in inflammation progression in some diseases such as HELLP syndrome.In another study, lncRNA mir-100-let-7a-2-mir-125b-1 cluster host gene (MIR100HG) inversely correlated with the expression of miR-204-5p and caused down-regulation of this marker in some malignancies (Huang et al. 2019).This lncRNA induced by Transforming Growth Factor b (TGFb) (Li et al. 2021).TGFb induces the release of pro-inflammatory cytokines including IL-1, IL-6 and TNFa and causes inflammation (Marek et al. 2002).Therefore, inhibition of this lncRNA may have a positive impact on HELLP syndrome alleviation.Moreover, the higher level of some inflammatory cytokines and chemokines were reported in HELLP syndrome samples.TNFa was identified as one of these cytokines.Overexpression of this factor along with some other inflammatory elements including highly sensitive C reactive protein (hsCRP) were reported in HELLP syndrome which can cause inflammation during pregnancy (Uckan and Sahin 2018).

TNFa
TNFa is the key pro-inflammatory factor that induce NF-jB signalling pathway.This pathway is a crucial one for inflammation process.Regarding to some studies, up-regulation of NF-jB p65 was also identified in patients with HELLP syndrome (Philip et al. 2004, Wang et al. 2021).Mounting studies indicated significant role of some lncRNAs in activating NF-jB pathway via promoting TNFa level.LncRNA HOX antisense intergenic RNA (HOTAIR) up-regulation was reported as a marker which leads to higher level of TNFa and can activate the NF-jB pathway (Wu et al. 2016, Emami et al. 2022a).This lncRNA regulates some up-stream regulatory factors of glucose metabolism including Hypoxia-inducible factor 1-alpha (HIF1a) and Phosphatase and tensin homolog (PTEN) indicating its function in glucose metabolism; thus, this lncRNA play a vital role in lipopolysaccharide-induced Glut1 expression via NF-jB pathway (Obaid et al. 2021).Hence, silencing of this lncRNA may be beneficial to prevent the inflammation process during HELLP disorder.Besides, the lncRNA myocardial infarction associated transcript 1 (Mirt1) has been illustrated to induce inflammatory cell infiltration via NF-jB pathway activation in some diseases.Knock down of Mirt1 could inhibit the NF-jB pathway and prevent the inflammatory factors in a variety of disorders including HELLP syndrome (Li et al. 2017b, Tutunchi et al. 2022).The interactions of some lncRNAs and miRNAs with NF-jB were also detected in some syndromes which regulate gene expression patterns at different facets.Some lncRNAs such as metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) can interact with NF-jB signalling pathway.This lncRNA may recruit some factors such as polycomb repressive complex 2 (PRC2) to the promoters of pro-inflammatory genes.Thus, MALAT1 can inhibit these factors epigenetically which may cause overexpression of inflammatory factors.It is also reported that MALAT1 transcript may directly linked to inflammatory transcripts to stimulate greater inflammatory response (Biswas et al. 2018, Liu et al. 2022a).Indeed, knock down of this lncRNA can inhibit this signalling pathways and therefore, suppress the inflammatory condition (Ghafouri-Fard et al. 2021) in a variety of disorders including HELLP syndrome.Some evidence also showed the role of mitogenactivated protein kinases (MAPKs) in the organogenesis of the placenta.P38 MAPK is the protein which is activated by some inflammatory cytokines including IL-1 and TNF-a and some other growth factors.Loss of p38 MAPK can lead to HELLP syndrome and placental defects (Triggianese et al. 2014).Some lncRNAs also have been reported as the factors that play a role in inhibition of inflammatory factors and p38 MAPK pathway.According to the study by Lu et al. lncRNA small nuclear RNA host gene 1 (SNHG1) is detected as a factor which prevents the expression of some inflammatory cytokines including TNF-a.Furthermore, this lncRNA inhibits the expression of miR-16-5p mediated factors which is associated with p38 MAPK signalling pathway.These data can demonstrate the positive effect of lncRNA SNHG1 in suppressing of inflammation which can be really important in alleviation of HELLP syndrome symptoms (Lei et al. 2019).LncRNA MALAT1 is also defined as a factor that increased the function of p38 MAPK pathway, indicating the reverse impact of this factor in inflammation process.Therefore, using si-MALAT1 can inhibit the p38 MAPK pathway and decrease inflammation (Fan et al. 2019).Regarding to all these studies, a variety of lncRNAs can play a role in a variety of disorders and the inflammation development or alleviation.Therefore, they are markers which their roles could be important in HELLP syndrome diagnosis and treatment.

LncRNAs and thrombosis
The thrombotic microangiopathy (TMA) is a disorder that represents some features such as disruption of the microvascular endothelium.HELLP syndrome has TMA features which are formed by microvascular endothelial injury events that cause microthrombi (Fang et al. 2008).Progression of many signalling pathways such as PI3K-AKT are involved in HELLP syndrome progression (Gong et al. 2021).Mounting studies indicate the role of some lncRNAs in the progression of thrombosis.In a study by Wei et el., lncRNA antisense noncoding RNA (ANRIL) can promote thrombosis.This lncRNA exerts its role via interacting with some factors in autophagy pathway.It causes beclin-1 overexpression as an important protein in autophagy pathway.ANRIL also inhibits the expression of miRNA-449a and miRNA-99a and leads to autophagy development (Zeng et al. 2019).Indeed, ANRIL can remove miR-99a and miR-499 during the autophagy process and then causes up-regulation of thrombomodulin and development of angiogenesis and thrombosis in human umbilical vein endothelial cells (Rey et al. 2021).Overexpression of Phosphatase and tensin homolog (PTEN) and Matrix Metallopeptidase 2 (MMP2) are associated with progression of Akt/autophagy signalling pathway which causes thrombosis development (Sun et al. 2019).In addition, the autophagy pathway is associated with megakaryocytic differentiation process.Overexpression of two autophagy markers including beclin-1and Microtubule-associated protein 1 A/1B-light chain 3 (LC3 II) increase megakaryocytic differentiation.There is also some evidence indicates that autophagy pathway is linked to many thrombopoietic markers such as thrombopoietin (TPO).This protein affects the majority steps of differentiation and maturation of megakaryopoiesis and thrombopoiesis.This protein interacts with thrombopoietin receptor (Mpl) and thereby activates JAK2 signalling, STAT3/5 and MAPK pathways which result in more production of megakaryocytes and platelets (Hill et al. 2020).In addition, some lncRNAs such as H19 also causes development of autophagy pathway via PI3K-AKT-mTOR signalling pathway (Ghafouri-Fard et al. 2022).This lncRNA activates this pathway by up-regulation of p70 S6 kinase (p70S6k) protein.This protein is the downstream substrate of mTORC1; therefore, activation of mTORC1 inhibits p70S6K and eIF4E-binding protein (4EBP1) and leads to autophagy prevention.Indeed, LncRNA H19 blockades the mTORC1 activation via p70S6K overexpression and subsequently activation of autophagy pathway (He et al. 2016, Xu et al. 2018).By activating the autophagy pathway via lncRNA H19, some cellular components are degraded by lipid oxidation.This process leads to loss of endothelial cells and subsequently development of advanced plaques, more platelet adhesion and thereby thrombosis progression (Yu et al. 2021).According to this data, lncRNA H19 may play a role in the formation of thrombosis in some diseases such as HELLP syndrome.In another study by Xinliang et al. lncRNA Silent information regulator 1 (Sirt1) results in up-regulation of Sirt1 protein and thereby Fork head box O3 (Foxo3a) degradation.Sirt1 is a marker which is linked to senescence and inhibits endothelial senescence as well as decreases Deep vein thrombosis (DVT).In addition, P53 as a downstream factor of Sirt1, is deacetylated via this protein.Therefore, the activity of P53 and some other downstream proteins including p21 and p16 also reduced and thereby activation of p53/p21 pathway is inhibited by Sirt1 overexpression.LncRNA Sirt1 can bind to Sirt1 3 0 UTR and causes more stability of this protein and its abundance at both mRNA and protein level (Lou et al. 2021, Yang et al. 2022).Thus, this lncRNA alleviates the formation of thrombus.This data can show the important role of lncRNA Sirt1 in reduction of thrombosis; therefore, it may be a vital biomarker in thrombosis alleviation during HELLP syndrome.According to some recent research, lncRNA GUSBP5-AS (enst00000511042) is increased in endothelial progenitor cells (EPCs) of DVT disorder.This lncRNA develops migration and angiogenesis of EPCs via modulating Fork head Box Protein O1 (FOXO1).Moreover, this lncRNA is associated with miR-223-3p and causes down-regulation of this miRNA which inhibits FOXO1.Overexpression of FOXO1 causes tissue factor (TF) development as well as thrombomodulin (TM) inhibition; therefore this factor results in thrombosis progression (Xie et al. 2020).There is also some evidence showed the role of lncRNA GUSBP5-AS in activation of Akt pathway by enhancing the expression of fibroblast growth factor 2 (FGF2), MMP2/9 and F-actin.Many research illustrates that overexpression of FGF2 increases the protein level of TF and subsequently development of thrombosis (Lee et al. 2011a).MMP2 and MMP9 are also platelet-associated markers and regulate the formation of acute thrombus markers and subsequently point limit thrombus formation via their effects on collagenolytic activity (Mastenbroek et al. 2015).This data shows the role of lncRNA GUSBP5-AS in thrombosis (Sun et al. 2020).and may be a novel marker for thrombosis therapy in a variety of diseases including HELLP syndrome.In another study by Yaming et.al., the role of lncRNA MALAT1 is identified in the progression of thrombosis.This lncRNA regulates the function of EPCs as well as Wnt/b-catenin signalling pathway.This signalling pathway is a downstream target of MALAT1; thus, any alteration in the biological function of EPCs, resulted from knock down of MALAT1, is reversed by prevention of the Wnt/b-catenin signalling pathway (Du et al. 2020).The Wnt axis is activated by Wnt ligand and then b-catenin phosphorylation is inhibited by glycogen synthase kinase-3b (GSK3b) at the N-terminus.This process demonstrates the stability of b-catenin destruction complex; therefore, this protein collects in the cytoplasm and then translocate into the nucleus.As a result, it can regulate the expression of Wnt target genes (Zhao et al. 2018).Indeed, MALAT1/Wnt/b-catenin axis may has an important role in the proliferation of EPCs and thereby progression of DVT (Du et al. 2020).Thus, lncRNA MALAT1 also may have a role in thrombosis development in patient with HELLP syndrome.Regarding to the study by Xiao-Qiang et. al., lncRNA Wilms tumour 1 associated protein pseudogene 1 (WTAPP1) positively regulates tube formation in EPCs via overexpression of MMP-1 and thereby activates the PI3K/Akt/mTOR pathway.Moreover, this lncRNA is a target for miR-3120-5P and it can be suppressed by this miRNA.This process results in down-regulation of MMP-1.Therefore, lncRNA WTAPP1 plays a significant role in the pathogenesis of DVT in various disorders including HELLP syndrome (Li et al. 2018b).All these previous studies illustrate the association of various lncRNAs with thrombosis; thus, it can be concluded that different kinds of lncRNAs may have a significant role in the progression of thrombosis in HELLP disorder and further studies about these biomarkers would be effective in treatment of HELLP syndrome.

Proliferation and autophagy
Severe HELLP syndrome is associated with vascular and villous lesions and it is also indicated that high placental dysfunction has occurred in this syndrome (Weiner et al. 2016).According to some studies various lncRNAs have a vital role in preeclampsia (PE) and placental dysfunction.In the study by Nan et.al., the lncRNA growth arrest associated lncRNA 1 (GASAL1) is linked to PE progression and downregulated in placentas in pregnant women with PE in comparison with healthy ones.Serine/arginine splicing factor 1 (SRSF1) is an RNA-binding protein of GASAL1.It is verified that these two markers have synergistic impacts on trophoblast cell invasion, proliferation, and apoptosis.There is also some evidence demonstrated that GASAL1 and SRSF1 exert their role via activating mTOR signalling pathway.these markers induce the expression of pmTOR and p4EBP1 and thereby activate the mTOR signalling pathway (Liu et al. 2020).The mTOR signalling pathway affects the use of some substrates which are involved in cell growth.This pathway can lead to the association of some signals from growth factors, extracellular matrix, and nutritional factors to improve the foetal growth.However, in extreme nutrient deficiency condition, the autophagy pathway is activated by mTOR pathway.It is also regulating L amino acid transport in placental villi and associates with hypoxia-induced autophagy in human trophoblast.Therefore, excessive autophagy eventually can cause placental dysfunction (Longtine andNelson 2011, Barahman et al. 2022).

Angiogenesis and apoptosis
In another study by Xu Li et. al., showed the lncRNA Xinactive specific transcript (Xist) is related to angiogenesis in placenta.Indeed, the level of this factor decreases in placenta and down-regulation of this lncRNA can cause a pathological condition in placental angiogenesis.In addition, the lncRNA-miRNA-mRNA regulation networks are shown to have a vital role in placental angiogenesis.LncRNA Xist regulates the expression of miR-429 via a molecular sponge.Vascular endothelial growth factor A (VEGF-A) is a target of miR-429 and its luciferase activity is decreased via this miRNA.Sponging of miR-429 by lncRNA Xist and suppressing of VEGF lead to abnormal placental angiogenesis.Therefore, Xist-miR-429-VEGF-A stimulates adverse foetal development (Chen et al. 2018).As a result, this lncRNA may be an important marker in placental dysfunction growth in a variety of syndromes including HELLP syndrome.There also exists some evidence illustrates the role of lncRNA NEAT1 in the progression of placental dysfunction.The mRNA level of this lncRNA is increased more than 4-fold in the villous trophoblasts of Intrauterine growth restriction (IUGR) placentas; thus, it results it placenta dysfunction (Gremlich et al. 2014).LncRNA NEAT1 causes the significant up-regulation of miR-411-5p and thereby inhibition of PTEN expression, as PTEN is the direct target for miR-411-5p.Thus this lncRNA leads to the promotion of trophoblast, migration and invasion in patients with PE (Fan et al. 2021).Moreover, lncRNA NEAT1 activates the cellular proliferation, migration, and invasion via PI3K/AKT/mTOR and MAPK extracellular-signal-regulated kinase (ERK) signalling as well as WNT/b-Catenin pathway (Katsushima et al. 2021, Jin et al. 2022).With this data, it can be concluded that lncRNA NEAT1 could be a cause of placental dysfunction in HELLP syndrome and may be a good target to treat placental dysfunction in this syndrome.Some research indicates the role of lncRNA HOXD-AS1 in the development of placenta tissue dysfunction via regulating MAPK pathway in pregnant women with PE.MAPK pathway is related to trophoblast function and included some important proteins such as p38 MAPK, JNK and ERK which are involved in regulating proliferation, apoptosis, differentiation and migration.Overexpression of lncRNA HOXD-AS1 is associated with reduction of p-p38 and p-JNK as well as up-regulation of p-ERK in MAPK signalling pathway in patient with PE which causes placental dysfunction (Jiang andZhao 2018, Emami et al. 2022b).In another study by Jing et.al., lncRNA Prospero homeobox 1-antisense RNA 1 (PROX1-AS1) expression is increased in both blood samples and placenta tissues of pregnant women with PE.Overexpression of this factor leads to PE via its interaction with caspase-9 and miR-211-5p.Indeed, lncRNA PROX1-AS1 exerts its role via sponging the miR-211-5p and thereby regulates the caspase-9.Elevation of caspase-9 increases the apoptosis rate and inhibits the cell migration and invasion by overexpression of miR-211-5p in the patients with PE.Although the apoptosis process is an essential step in placental development, the overactivation of this process causes placental diseases.Caspase-9 is activated via cytochrome c in cytosol.This factor is bound by apoptosis protease and causes the activation of factor-1 (APAF-1) as well as formation of apoptosome.Apoptosome causes cleavage of caspase-9 and activation of this factor.By activating of caspase-9, the terminal apoptosis pathway is activated (Sharp et al. 2010).With these results, it can be concluded that silencing lncRNA PROX-AS1 might be beneficial to alleviate the PE progression (Tang et al. 2021) and it may also be a good biomarker to detect some disorders such as HELLP syndrome.According to the association of multiple lncRNAs with placental dysfunction, investigating the role of these markers in the development of placental dysfunction in patients with HELLP syndrome might help to find some novel ways for HELLP syndrome alleviation.

Treatment approach for HELLP syndrome
The treatment of HELLP syndrome is different based on the type of pathogenesis and the condition of the patients, however, it has been determined that therapeutic approaches based on lncRNAs in patients can be effective in improving them to some extent (Bazzan et al. 2020).Recently, the use of therapeutic approaches based on targeting lncRNAs has been used in many diseases, however, this type of treatment has not been commonly performed in HELLP syndrome.
Based on studies, it has been shown that the use of dexamethasone in HELLP syndrome patients can be effective in treating hypertension and preventing the progression of the disease.On the other hand, it has been determined that lncRNA MALAT1 can play a role in hypertension through the synthesis of VEGF (Maghsoudi et al. 2022).Therefore, it can be said that the use of dexamethasone can play a role in the recovery of HELLP syndrome patients by controlling the expression of MALAT1 (Li et al. 2017a;Fonseca et al. 2019).Inflammation is another factor of pathogenesis in HELLP syndrome.Some lncRNAs cause disruption in endothelial cells and the development of HELLP syndrome by producing inflammatory mediators.Therefore, targeting them can be considered as a treatment option (Table 1).

Conclusion
HELLP syndrome is a life-threatening disorder which its diagnosis and treatment is vital during pregnancy.In this review, we investigated the role of some lncRNAs in the pathogenesis of HELLP syndrome.Some of these markers cause the progression of the disease; however, others alleviate the symptoms and thereby inhibit the development of this syndrome.Therefore, detection of these lncRNAs and the related signalling pathways as well as some down-stream elements such as miRNAs that are identified as important regulators of lncRNAs, might be beneficial to find novel strategies for prevention and treatment of patients with HELLP syndrome.

Disclosure statement
No potential conflict of interest was reported by the author(s).Leads to the overexpression of lncRNA LOC10369133 (Martin Jr et al. 2012, Wallace et al. 2013, LaMarca et al. 2011, Li et al. 2019, Liu et al. 2022b) Glyceryl trinitrate Leads to lower maternal blood pressure via improving aortic compliance Leads to the overexpression of lncRNA A930013F10Rik (Cetin et al. 2004, Jeong et al. 2018, Johal et al. 2014, ENOS Trial Investigators 2015) Nifedipine Developing renal blood flow and resulting in reduction of vascular resistance as well as increasing urine output via inhibition of antidiuretic hormones and Ca2þ channel - (Xiao et al. 2017, O'Brien andBarton 2005) Dexamethasone Decadron Used for steroid therapy for foetal lung maturity during HELLP syndrome via increasing the activity of choline phosphate cytidylyltransferase Causes the higher expression of lncRNA ZNF197-AS1 (Zhang et al. 2021, Ferguson et al. 2009, Gross et al. 1980) PVT1: Plasmacytoma variant translocation 1, TF: tissue factor, NF-jB: nuclear factor kappa light chain B cells, AP-1: activator protein 1, MCP-1: monocyte chemotactic protein, TNF-a: tumour necrosis factor enhancer of activated alpha, A930013F10Rik: RIKEN cDNA A930013F10 gene, ZNF197-AS1: zinc finger protein 197 antisense RNA 1 (Table 2).

Funding
The author(s) reported there is no funding associated with the work featured in this article.

Table 1 .
Summary of drugs used for HELLP syndrome patients.