Novel Insights into Prokineticin 1 Role in Pregnancy-related Diseases

Abstract

Prokineticin 1 (PROK1) is a secreted protein involved in a range of physiological activities such as cell proliferation, migration, angiogenesis, and neuronal cell proliferation. Emerging evidences show that PROK1/PROK receptors (PROKRs) are expressed by trophoblasts, and decidual stroma cells at the maternal-fetal interface. PROK1 plays a critical role in successful pregnancy establishment by regulating the decidualization, implantation and placental development. Dysregulation of prokineticin signaling has been described in certain pathological states associated with pregnancy, including pre-eclampsia, recurrent miscarriage and fetal growth restriction. In this review, the expression and pleiotropic roles of PROK1 under physiological and pathological pregnancy conditions are discussed.

1. Introduction

Prokineticin family include two members, prokineticin 1 (PROK1; also called EG-VEGF) and prokineticin 2 (PROK2; also called Bv8). The Human PROK1 gene maps to the regions of chromosome 1p13.1 and is composed of three exons with no alternative splicing product. The mature PROK1 protein is formulated of 86 amino acids and its relative molecular weight is 8.6 kDa ¹. The Human PROK2 gene maps to chromosome 3p21.1 ² and encodes a mature protein of 81 amino acid ¹. PROKs are highly conserved in varies species, characterized by a conserved N-terminal sequence (AVITGA), which is essential for the correct binding of receptors ³.

These two secreted proteins act through two G protein-coupled receptors, prokineticin receptor 1 (PROKR1) and prokineticin receptor 2 (PROKR2) ⁴. PROKR1 is mainly involved in cell proliferation and angiogenesis, while PROKR2 is involved in the vital vascular endothelial permeability process. Receptors couple to Gq ⁵, Gi ⁶ and Gs ⁷ protein. Prokineticin receptors (PROKRs) mediate the intracellular calcium mobilization, phosphorylation of p44/p42 mitogen-activated protein kinase, serine/threonine kinase Akt and cyclic AMP (cAMP) accumulation, respectively ⁷. Additionally, PROKRs can either stimulate or inhibit the accumulation of cyclic AMP (cAMP) via Gs or Gi proteins ^{7, 8}, respectively. Moreover, PROKRs have been shown to act through Gi protein to stimulate mitogen-activated protein kinase (MAPK) ⁶.

The PROK1 and PROK2 proteins are expressed in multiple organs and tissues, such as brain, heart, digestive tract, bone marrow, ovary, testis, decidua, and placenta. It was found that prokineticins showed dynamic expression throughout the physiological process in many organs, including circadian rhythm ⁹, menstrual cycle ¹⁰ and pregnancy ¹¹. Additionally, prokineticins are involved in the cancer formation¹².

The expression of prokineticin 1 in the reproductive system plays an important role in the development of the placenta. Prokineticin 1 could affect the growth and development of the placenta, including the processes of implantation ¹³, invasion and angiogenesis ¹¹. Whereas the prokineticin 2 mainly affected in the central nervous system and could cause idiopathic hypogonadotropic hypogonadism by abnormal expression. The function of PROK2 in pregnancy-related disease is unclear and needed further study. In pregnancy-related diseases, PROK1 is related to preeclampsia ¹⁴, recurrent miscarriage ¹⁵, intrauterine growth restriction ¹⁴, premature birth ¹⁶ and other diseases. In this article, we are going to discuss the role of PROK1 in normal pregnancy and pregnant-related diseases.

2. Biological functions of the prokineticin1

Through constant study, researchers have identified many physiological functions of PROK1. The involvement of PROK1 in diverse physiological activities further demonstrates its important research value.

2.1 Prokineticin1 in ovary

Ovary is one of the organs where PROK1 protein highly expressed. Researchers discovered that PROK1 was expressed in luteal steroidogenic cells of human ovaries. Luteal endothelial cells (LEC) expressed high levels of both PROKR1 and PROKR2 protein. PROK1 protein enhanced proliferation of luteal endothelial cells by increasing [3H]-thymidine binding, MAPK activation and c-jun/fos mRNA expression. The expression of PROKR2 protein is increased in LEC under some stress conditions,such as tumor necrosis factor α (TNFα) and chemical hypoxia,while the PROKR1 protein expression level is unchanged ¹⁷. Exogenous PROK1 protein can inhibit LEC apoptosis ¹⁷.

This implies that PROK1 may exert an anti-apoptotic effect on LEC through PROKR2. In addition, PROK1 increased VEGF mRNA expression in bovine luteinizing steroid-producing cells via PROKR1 ¹⁷. These findings suggest that PROK1 plays an important role in luteal function by promoting LEC proliferation and anti-apoptotic effects ¹⁸. Researchers compared PROK1 expression in 13 pairs of human polycystic ovary syndrome (PCOS) and normal ovary samples. PROK1 was highly expressed in theca interna and stroma of PCOS ovaries, suggesting that the angiogenic function of PROK1 may be related to the cyst formation ¹⁹. In another study, researchers transfected rat ovarian granule cells with miR-28-5p mimics, and found that PROK1 protein level is downregulated ²⁰. MiR-28-5p reverses the promotive effect of PROK1 on cell proliferation and the inhibitory effect on autophagy ²⁰. PROK1 promoted proliferation and inhibited the apoptosis of rat ovarian granulosa cells through the AKT/mTOR signaling pathway ²⁰.

Additionally, concentration of PROK1 protein in the follicular fluid is relevant to the follicular size and is predictive of the oocyte competence ²¹. Follicle Stimulating Hormone and hCG up-regulated PROK1 protein secretion in cumulus cell primary cultures, probably through the cAMP pathway ²¹. Successful embryo implantation may be predicted by PROK1 concentration in the follicular fluid and fertilization culture media in conventional in vitro fertilization-embryo transfer ²². Moreover, expression of PROK1 and PROK2 is correlative to the germ cell development in the human fetal ovary ²³. PROK1 increases ERK phosphorylation and COX2 expression ²³.

2.2 Prokineticin1 in gastrointestinal tract

PROK1 protein was originally identified as a promoter of gastrointestinal smooth muscle contraction ²⁴. PROK1 has a high-affinity site in human ileal smooth muscle ²⁴. Additionally, PROKR1 mRNA is highly expressed in rat ileum ²⁵. In rat ileum, PROK1 induces a biphasic contractile response, including an early tetrodotoxin (TTX)-sensitive response and a late TTX-insensitive response ²⁵.

PROK1 mRNA is not expressed in the normal colorectal mucosa, but was detected in all colorectal cancer cell lines ²⁶. Researchers found that PROK1 increased the microvascular density in colon cancer and promoted cancer cell proliferation ²⁶. It also increased the liver metastatic ratio ²⁶. A recent study showed that PROK1 was involved in lymphatic vessel production and participated in lymphatic metastatic pathways ²⁷.

2.3 Prokineticin1 in the bone marrow and blood cells

In human or mouse hematopoietic stem cell cultures, PROK1 increases the number of granulocytic and monocytic colony-forming units ²⁸. Systemic in vivo exposure to PROK1 significantly increased total leukocytes, neutrophils and monocytes ²⁸. Moreover, PROK1 could protect multiple myeloma (MM) cells from apoptosis by upregulated Myeloid-cell-leukemia 1 (Mcl-1) ²⁹. Furthermore, Mcl-1 was upregulated through MAPK, AKT and STAT3 pathway ²⁹.

2.4 Prokineticin1 in neural crest cells

PROKR1 and PROKR2 are expressed in the human enteric neural crest cells (NCCs) ^{30, 31}. PROK1 protein activates the Akt and MAPK pathways and induces proliferation and differentiation of NCCs via PROKR1 ³⁰. Neuroblastoma is derived from improperly differentiated neural crest cells. PROK1 induces neuroblastoma cell proliferation by activating the Akt pathway through PROKR1 and PROKR2 ³². PROKR2 is essential for inhibiting neuroblastoma cell apoptosis ³².

3. The expression of PROK1 at the maternal-fetal interface

3.1 Endometrium

Immunohistochemistry and in situ hybridization analysis showed that PROK1, PROKR1 and PROKR2 were located in a variety of cellular components of the human uterus, including the glandular epithelium, stroma and endothelial cells of the endometrium, as well as in smooth muscle and endothelial cells of the myometrium ³³ (Table 1). Immunohistochemistry analysis showed that PROK1 was expressed primarily in the glandular epithelial cells and that its expression was dynamic during the menstrual cycle, and reached a peak in expression during the secretory phase³⁴. PROK1 mRNA expression in endometrial cells is controlled by E2 (estrogen) and P4 (progesterone) ³⁴. Additionally, PROK1 was rarely detected in the endometrial samples from the postmenopausal patients ³⁴. This suggests that PROK1 expression is highly correlated with steroid hormones.

Table 1.

The expression of PROK1, PROKR1, and PROKR2 at maternal-fetal interface

Site	Cell	PROK1	PROKR1	PROKR2	Reference
Endometrium	Glandular epithelial cell	+*	+	Weak	26
	Stromal cell	+	+	-	26
	Endothelial cell in the endometrium or Myometrium	+	+	Weak	26
	Smooth muscle cell	+	+	-	26
Decidua	Decidual cell	+	+	+	35
Placenta	Syncytiotrophoblast	+	+	+	10,37
	Cytotrophoblast	+	+	Weak	10,36
	Extravillous trophoblast		Weak	+	10
	Placental microvascular endothelial cells	+	+		36
	Placental endothelial cells		-	+	10
	Macrophages		+	+	10,36

*: PROK1 is expressed primarily in the glandular epithelial cells.

3.2 Decidua

The researchers found that PROK1 and its receptors PROKR1 and PROKR2 were expressed in decidual cells and play an important role in the process of endometrial decidualization ³⁵. Expression of PROK1 mRNA in human first-trimester decidua was significantly increased compared to non-pregnant endometrium ³⁵.

3.3 Trophoblasts

The expression of PROK1 mRNA and PROKR1 mRNA peaks in trophoblast cells at 8-10 weeks of gestation ³⁶. PROK1 protein is localized to the syncytiotrophoblasts ³⁶, expressed minimally in cytotrophoblasts and not expressed in the extravillous trophoblasts ³⁷. PROKR2 mRNA is not expressed in trophoblasts in the first trimester of pregnancy, except for low levels of expression detected at 8-10 weeks of gestation ^{37, 38}. Moreover, PROKR1 protein is highly expressed in cytotrophoblasts, placental microvascular endothelial cells and Hofbauer cells, whereas PROKR2 protein is mainly expressed in syncytiotrophoblasts, Hofbauer cells and extravillous trophoblasts ³⁷.

4. Regulation of prokineticin1 expression at maternal-fetal interface

4.1 Hormone

Researchers found that human chorionic gonadotropin (hCG) could induce the expression of PROK1 in a baboon model, human endometrial epithelial cells ³⁹ and first-trimester decidua ³⁹ (Table 2). They showed by dual immunohistochemical analysis that hCG receptors, PROK1 and PROKR1 were present in the glandular epithelial cells of the first trimester decidua ⁴⁰. PROK1 and hCG show similar expression pattern during the first trimester of pregnancy. PROK1 expression peaks in trophoblast cells at 8-10 weeks of gestation, while the highest values of hCG are shown at 10-11 weeks of gestation³⁹. Human chorionic gonadotropin increases the expression of PROK1, PROKR1 and PROKR2 mRNA and protein in dose- and time-dependent manners³⁹. In another study, researchers found that primary endometrial stromal cells decidualization by progesterone and cyclic adenosine monophosphate could be inhibited by PROK1 knockdown³⁹. It is reported that hCG regulated PROK1 expression in the ovary ⁴¹. HCG can increase cAMP levels, and cAMP enhances PROK1 promoter expression³⁹.

Table 2.

Regulation the expression of PROK1, PROKR1, and PROKR2 at maternal-fetal interface

Hormone	PROK1	PROKR1	PROKR2	Reference
Human chorionic gonadotropin	+	+	+	39
Progesterone	+	+		11,26,34
Estrogen	+	+		42
Dihydrotestosterone*	+			44
Insulin	+			43
Hypoxia	+	+		37
PPARγ	+	+	+	47

* Only effect with insulin.

Battersby et al. ³³ found that during menstrual cycle the expression of PROK1 protein in the endometrium showed periodic changes. The PROK1 protein showed higher expression in the secretory phase of the menstrual cycle than that in the proliferative phase ³³. In endometrial cell culture, PROK1 mRNA was detected in endometrial cells only in the presence of steroids ³⁴. Using the porcine model of intrauterine infusions of estradiol-17 beta, researchers revealed that it upregulated endometrial expression of PROK1 and PROKR1 mRNA. Moreover, estradiol-17 beta, acting together with prostaglandin E2 (PGE2) increases PROKR1 protein expression in the endometrium ⁴².

Ujvary et al. ⁴³ found that insulin upregulated PROK1 in a dose-dependent manner in decidualizing endometrial stromal cells. Furthermore, dihydrotestosterone is demonstrated to upregulate both of gene and protein expression of PROK1 along with insulin. But dihydrotestosterone alone do not show significant effects at all ⁴⁴. On the one hand, insulin interacting with dihydrotestosterone enhances the decidualization, but on the other hand dysregulates the migration of decidual cells and the invasion of trophoblast cells ⁴⁵.

4.2 Hypoxia

It was reported that hypoxia-inducible factor 1-alpha (HIF1α) response element was identified in the promoter region of PROK1 ⁴⁶. Researchers confirmed that PROK1 mRNA and PROKR1 mRNA were upregulated by hypoxia in the human placenta ³⁷. Additionally, the stimulating effect of insulin on PROK1 gene and protein expression is mediated by HIF1α acting through the phosphatidylinositol 3-kinase (PI3K) signaling pathway ⁴³.

4.3 Peroxisome proliferator-activated receptor-γ

Researchers confirmed that upregulating peroxisome proliferator-activated receptor-γ (PPARγ) increased the PROK1 and its receptors expression in both mRNA and protein level. On the other hand, the PPARγ^-/- mice showed low level expression of PROK1 ⁴⁷. It is shown that PPARγ plays an important role in the control of trophoblast migration and invasion ⁴⁸. It was found that PPARγ upregulated PROK1 and inhibited trophoblast invasion via PROKR2 ⁴⁷.

5. Role of prokineticin1 in normal pregnancy

5.1 Regulation of trophoblast functions

The depth of invasion of placental trophoblast cells into the uterine wall is quite important in normal pregnancy. Therefore, during the placental development, the proliferation, differentiation and invasion of trophoblast cells are finely regulated. Diverse growth factors and cytokines stimulate differentiation of trophoblast cells to an invasive phenotype, such as epidermal growth factor (EGF), hepatocyte growth factor (HGF), transforming growth factor beta (TGF-β), insulin-like growth factor-II (IGF-II), and interleukin-1 (IL-1) ⁴⁹.

Several studies have confirmed that PROK1 played an important role in trophoblast invasion, affecting trophoblast proliferation, migration, and invasion (Figure 1). Extravillous trophoblastic cells have an important role in the establishment of a successful pregnancy. These cells invade the decidua and spiral arteries to establish the fetal-mother circulation. PROK1 enhanced the invasion of extravillous trophoblastic cells (EVTs) by upregulating the expression of matrix metalloproteinases (MMP) 2 and MMP9 mRNA. And this upregulation was achieved through the receptor PROKR2 ⁵⁰. They also found the upregulation of extracellular signal-regulated kinase (ERK) 1/2 after the treatment of high concentration of PROK1 ⁵⁰. Researchers found that PROK1 activated ERK1/2 signaling and subsequent upregulation of MMP2 and MMP9 mRNA levels, thereby stimulating invasion of human trophoblast HTR-8/Svneo cells ⁵¹. Therefore, the decrease in PROK1 circulating and placental levels at the end of the first trimester may, with other factors, contribute to EVT invasion and to the establishment of fetal-maternal circulation ⁵².

Figure 1.

Role of prokineticin1 in the maternal-fetal interface. PROK is secreted mainly by placenta. PROK1 can promote proliferation, differentiation, migration and invasion of trophoblasts through upregulation of MMPs and other pathways. PROK1 acts on placental microvascular endothelium and affects angiogenesis. It acts on endothelial stromal cells and affects decidualization through upregulation of COX2, dickkopf1, etc. PROK1 affects placental implantation through regulation of IL8, IL11 and other cytokines. By Figdraw (www.figdraw.com).

Primary cilia are cellular antennae that receive environmental signals and are essential for normal development. PROK1 and its receptors were detected in primary cilia ⁵¹. It was found that inhibition of primary cilia growth could cause decrease of PROK1 and MMP9 expression ⁵¹. In a study published recently, researchers showed that miR-346 and miR-582-3p regulated PROK1-induced trophoblast invasion through repressing MMP 2 and MMP 9⁵³. It was shown that miR-346 and miR-582-3p inhibited not only PROK1 expression but also trophoblast invasion and migration in JAR and HTR-8/Svneo cell lines⁵³. The PROK1-stimulated cell proliferation was mediated by PI3K/AKT/mTOR, MAPK, and cAMP ¹³. PROK1 enhanced adhesion of trophoblast cells to fibronectin and laminin matrices, which are mediated predominantly via leukemia inhibitory factor (LIF) induction ⁴⁰.

5.2 Effect on endometrium/decidua

Decidualization is a complex process. Endometrial stromal cells are regulated by hormones as well as cytokines and undergo morphological, functional and genetic changes to support embryo implantation and development ⁵⁴. PROK1 and its receptors regulate the implantation and decidualization via affecting the expression of implantation-related genes. It has been shown that PROK1 can regulate the expression of many genes, including: LIF, COX-2, IL-6, IL8, and IL-11 ^{35, 42}. These cytokines are verified to be related to the implantation and decidualization.

Differentiation of endometrial stromal cells into decidua is mediated by PGE2 synthesis through elevation of COX-2 protein ^{35, 55}. In an experimentation conducted by Sharon et al, the expression of COX-2 protein was significantly increased after the treatment of PROK1 in a human germ cell tumor line. And it was confirmed that PROK1 signaling via PROKR1 induce the expression of COX-2 protein²³. Another research showed that PROK1 induced COX-2 expression and prostaglandin synthesis in human endometrial cells and first trimester decidua via a Gq coupled pathway ⁵⁶. Another study found that PROK1 could upregulate the Dickkocf1 which regulate cells proliferation and decidualization of endometrial stromal cells. It was confirmed through a Gq-calcium-calcineurin-nuclear factor of activated T-cells-mediated pathway ⁵⁷.

IL-8 is highly expressed in the endometrium and decidua and involves in several processes of endometrial physiology such as angiogenesis, proliferation, chemotaxis, trophoblast invasion and uterine contraction. Researchers found that in endometrial cells PROK1 via PROKR1 could activate the calcineurin/NFAT pathway to induce IL-8 expression ⁵⁸.

In human endometrial stromal cells, IL-11 has been shown to promote progesterone-induced decidualization, which implies a role for IL-11 in preparing the endometrium for implantation ⁵⁹. PROK1-PROKR1 induces the expression of IL-11 in Ishikawa cells and first trimester decidua in a guanine nucleotide-binding protein (G(q/11)), extracellular signal-regulated kinases, Ca²⁺ and calcineurin-nuclear factor of activated T cells dependent manner through the calcium-calcineurin signaling pathway ⁶⁰.

Connective tissue growth factor is considered to be up-regulated by PROK1 in the first-trimester decidua⁶¹. PROK1 regulates the expression of connective tissue growth factor by activating the Gq, PLC, cSrc, EGFR, MAPK/ERK kinase pathway⁶¹.

5.3 Effect on angiogenesis

Several studies have elucidated the angiogenic role of PROK1 in reproductive organs. Study found that PROK1 mainly acted on human placental microvascular endothelial cells (HPEC), and stimulated HPEC's proliferation, migration and survival, while VEGF mainly affected the umbilical vein-derived macrovascular endothelial cells ⁶². Compared to VEGF, PROK1 was more effective in stimulating HPEC sprout formation, pseudovascular organization, and it significantly promoted HPEC permeability and paracellular transport ⁶². In another study, researchers demonstrated that PROK1 controlled homeobox genes expression in normal human placenta and in placenta from fetal growth restriction (FGR) pregnancies ⁶³. The homeobox genes expressed in placental microvascular endothelial cells may play a role in the regulation of epithelial-mesenchymal interactions in the placenta and proliferative capacity of placental microvascular endothelial cells ⁶⁴.

The effects of PROK1 on HPECs suggests that this factor is involved in placental angiogenesis, a process closely related to placental growth and fetal maternal exchanges.

6. Role of prokineticin1 in pathological pregnancy

The dynamic expression of PROK1 throughout pregnancy, its multiple roles in the villi development, and its regulation by hypoxia suggest that this cytokine contributes to the successful pregnancy establishment. Thus, inappropriate expression and/or dysfunction of PROK1 may lead to major complications of pregnancy (Table 3), such as recurrent implantation failure, recurrent pregnancy loss (RPL), ectopic pregnancy (EP), fetal growth restriction (FGR), choriocarcinoma, and pre-eclampsia (PE). These complications are associated with abnormal angiogenesis, as well as trophoblast invasion failures and abnormal placental development.

Table 3.

The role of PROK1 in pregnant related disease

Disease	Abnormal expression of PROK1	Functions of PROK1	Reference
Pre-eclampsia	higher	Inhibit trophoblast invasion	10,66, 67
Recurrent miscarriage	lower	gene polymorphisms of PROK1 and PROKR1 are related to recurrent miscarriage	15,70,71
Recurrent implantation failure	higher	unclear	72
Fetal growth restriction	high	Increase the angiogenesis and proliferation of placental villi	73

6.1 Pre-eclampsia

Pre-eclampsia is the most common hypertensive disease during pregnancy. There are several theories about the pathogenesis of pre-eclampsia. Although the etiology of PE is far from being fully understood, its development is thought to be primarily due to superficial invasion of the maternal decidua and spiral arteries by EVTs ⁶⁵. It creates a prolonged hypoxic environment in the placenta, and leads to the release of antiangiogenic factors from the placenta ⁶⁵. More specific and earlier markers associated with the disease need to be further investigated. PROK1 satisfies many of the characteristics of an early PE markers. It is abundant in the human placenta during early pregnancy, its expression is elevated by hypoxia, it controls trophoblast invasion, and has specific effects on placental endothelial cells. More importantly, PROK1 inhibit the EVTs which may play a critical role in the cause of PE.

Scientists found that the RNA and protein expression levels of PROK1 were significantly elevated in PE patients compared with age-matched control ¹⁰. The RNA and protein expression of PROK1 in the placenta of PE patients decreases significantly, suggesting that the in-situ expression of PROK1 in the placenta of PE patients may be down regulated at the transcriptional level ⁶⁶. Moreover, the RNA and protein expression level of PROKR1 shows no difference between control and PE patients ⁶⁶.

A group created a new mouse model of gestational hypertension by continuous injection of prokineticin1 ¹⁴. Sustained high levels of ghrelin over the first trimester of pregnancy significantly increased blood pressure and renal impairment in mice ¹⁴. It significantly decreased the thickness of the decidual and junctional zone in the placenta ¹⁴. Moreover, sustained high levels of prokineticin1 in late pregnancy reduced blood pressure in mice compared to mice injected only up to mid-gestation ¹⁴.

Both of the RNA and protein expression of PROK1 is upregulated by hypoxia. The promoter regions of the gene contain functional hypoxia response elements that bind to hypoxia-inducible factor 1 (HIF-1) and mediate its expression under hypoxic tension. PROK1 upregulation by hypoxia confirms its role in the first trimester, as placental development and angiogenesis occur in hypoxic environment during this period ⁶⁷.

The miRNA-200 family is predicted to target the PROK1 5'-untranslated region ⁶⁸. Increased miRNA-141 and miRNA-200a inhibit the protein expression of PROK1, matrix metalloproteinase 9 (MMP9) and downstream extracellular signal-regulated kinase (ERK) signaling, thus leading to trophoblast invasion failures ⁶⁸. The growth of the primary cilium is inhibited by increases in miR-141 and miR-200a expression ⁶⁸. The number of cilia in the human placenta of preeclampsia women was markedly decreased compared to normal placenta ⁶⁸.

6.2 Recurrent miscarriage

Recurrent miscarriage is related to abnormal blood vessels in the placental bed, suggesting that early disturbance of placental blood vessel development may lead to miscarriage ⁶⁹.

In 2010, Su et al. ¹⁵ studied the genes of 115 women with recurrent miscarriage and 170 women from the control group, and selected 11 single nucleotide polymorphisms from PROK1, PROKR1, and PROKR2 by using genotype analysis. Further using multifactor dimensionality reduction (MDR) method for analysis, they found that 2 SNPs of PROKR1 (rs4627609, rs6731838) and 1 SNP of PROKR2 (rs6053283) are closely related to recurrent miscarriage (P<0.05) ¹⁵. At the same time, the frequency of the C-G and T-A haplotypes of PROKR1 and the A-G-C-G-G haplotypes of PROKR2 were also significantly increased in recurrent miscarriage (P <0.05) ¹⁵.

Hence, the gene polymorphism and haplotype of PROK1 receptor are both related to recurrent miscarriage. In a study carried out in 2014, researchers found the PROKR1 (I379V) and PROKR2 (V331M) were nonsynonymous variants showed significant association with recurrent miscarriage ⁷⁰. It has been shown that the common variant of V67I may act as a modifier in the PROK1-PROKR system through down-regulation of PROK1 expression ⁷¹. PROK1 (V67I) has been shown to play a role as a modifier gene in the PROK1-PROKR system of human early pregnancy ⁷¹.

6.3 Recurrent implantation failure

In a recent study, researchers studied the endometrial samples of 15 women with recurrent implantation failure (RIF) and 15 women from control group. The results showed that PROK1 mRNA levels were 6.09 times higher compared to endometrial samples obtained from women with RIF than in samples obtained from women from control group, whereas PROKR1 mRNA levels were 2.46 times lower in endometrial samples obtained from women with RIF than in samples from women from control group. There was no statistically significant difference between women with RIF and women from control group regarding PROK2 and PROKR2 levels ⁷². The results suggested that expression of the PROK1/PROKR1 system could play an important role in implantation.

6.4 Fetal growth restriction

During pregnancy, fetal optimal growth depends on an adequate blood vascular network in the fetal part. Abnormal placental angiogenesis will endanger the supply of nutrients and hormones, which will eventually lead to FGR. Although the exact cause of FGR in not clear yet, several researches shows that the PROK1 and its receptors are highly associated with FGR. Scientists found that in FGR patients in the third trimester of pregnancy, both RNA and protein levels of PROK1 are significantly increased in the placenta ⁷³. Scientists found that PROK1, PROKR1, and PROKR2 mRNA and protein levels were significantly increased in FGR placentas ⁷³. PROK1 can increase the angiogenesis of placental villi and the expression of CD31, a marker of endothelial cells in the placental villi. At the same time, PROK1 promotes the proliferation of villi and the anchoring of cytotrophoblast layers, and promotes the survival of placental villi under stress conditions ⁷³. Therefore, the increase of PROK1, PROKR1 and PROKR2 occurs in FGR as a compensatory mechanism to ensure proper pregnancy progression.

6.5 Choriocarcinoma

It has been suggested that the development and progression of choriocarcinoma is associated with an over-proliferation of the trophoblastic layer, resulting in an increased mass of cells that acquire a migratory and/or aggressive phenotype ⁷⁴. Prokineticin 1 is demonstrated to upregulate trophoblast proliferation and invasion ⁷³. Prokineticin 1 protein and its two receptors are upregulated in the blood and placenta of choriocarcinoma patients ⁷⁵. Additionally, in the mouse model of choriocarcinoma, the application of PROKR1 or PROKR2 antagonists to gravid mice delayed tumor progression and increased the duration of pregnancy maintenance ⁷⁵.

7. Conclusions

During the past 20 years, the prokineticin system has been reported to play a role in a range of physiological functions. Mutations or dysfunction of these factors are also associated with a number of diseases. Targeting prokineticin system may provide treatments for diseases such as cancer, inflammation, pain, and neuropathology. In recent years, more and more concerns have been expressed about their specific role at the maternal-fetal interface. Since the PROK2 is mainly expressed in the central nervous system and PROK1 is widely detected in the steroidogenic organs, researches mainly focused on the PROK1 and further research is needed to reveal the functions of PROK2 in the maternal-fetal interface.

PROK1 is co-regulated by pregnancy-associated hormones, cytokines and other factors. It is highly expressed in embryonic trophoblasts and decidual stromal cells at the maternal-fetal interface, and participates in a series of physiological processes through interaction with its receptors, PROKR1 and PROKR2. PROK1 not only promotes the endometrial decidualization process and enhances uterine receptivity, but also induces migration and invasion of trophoblasts and facilitate embryo implantation and development. Nevertheless, abnormal expression of PROK1 may lead to impaired endometrial decidualization, trophoblastic dysfunction and placental dysplasia, resulting in pregnancy complications such as recurrent miscarriage, PE and FGR, although the exact mechanisms have not been fully clarified. Therefore, normal expression of PROK1 in the first trimester is essential for a successful pregnancy. Regulation of PROK1, PROKR1 and PROKR2 expression may be potential targets for clinical therapy of patients with abnormal pregnancies, although further studies are needed to confirm this.

Abbreviations

PROK1

prokineticin1

PROKR

prokineticin receptor

EG-VEGF

endocrine gland-derived vascular endothelial growth factor

cAMP

cyclic AMP

MAPK

mitogen-activated protein kinase

AKT

proto-oncogene protein c-akt

LEC

luteal endothelial cells

TNFα

tumor necrosis factor α

PCOS

polycystic ovary syndrome

Mcl-1

Myeloid-cell-leukemia 1

MM

multiple myeloma

hCG

human chorionic gonadotropin

PGE2

prostaglandin E2

HIF1α

hypoxia-inducible factor 1-alpha

PI3K

phosphatidylinositol 3-kinase

PPARγ

peroxisome proliferator-activated receptor-γ

EGF

epidermal growth factor

HGF

hepatocyte growth factor

TGF-β

transforming growth factor beta

IGF-II

insulin-like growth factor-II

IL-1

interleukin-1

EVTs

extravillous trophoblastic cells

MMP

matrix metalloproteinases

ERK

extracellular signal-regulated kinase

LIF

leukemia inhibitory factor

G(q/11)

guanine nucleotide-binding protein

HPEC

human placental microvascular endothelial cells

RPL

recurrent pregnancy loss

EP

ectopic pregnancy

FGR

fetal growth restriction

PE

pre-eclampsia

RIF

recurrent implantation failure