Porcine Placenta Research Articles

Effects of Ethanol Addition on the Efficiency of Subcritical Water Extraction of Proteins and Amino Acids from Porcine Placenta.

In a previous study, hydrolysates of porcine placenta were obtained and the extraction efficiency for proteins and amino acids was compared between sub- and super-critical water extraction systems; optimum efficiency was found to be achieved using subcritical water (170℃, 10 bar). In this study, the effects of adding ethanol to the subcritical water system were investigated. The lowest-molecular-weight extraction product detected weighed 434 Da, and the efficiency of extraction for low-molecular-weight products was increased when either the concentration of ethanol was decreased, or the extraction time was lengthened from 10 min to 30 min. The highest concentration of free amino acids (approximately 8 mM) was observed following 30 min extraction using pure distilled water. The concentration of free amino acids was significantly lower when ethanol was added or a shorter extraction time was used (p<0.05). Color change of the solution following extraction was measured. There were no significant differences in color between lysates produced with different extraction times when using distilled water (p>0.05); however, using different extraction times produced significant differences in color when using 20% or 50% ethanol solution for subcritical extraction (p<0.05). The range of pH for the hydrolysate solutions was 6.4-7.5. In conclusion, the investigated extraction system was successful in the extraction of ≤ 500 Da hydrolysates from porcine placenta, but addition of ethanol did not yield higher production of low-molecular-weight hydrolysates than that achieved by DW alone.

Effect of Porcine Placenta Extract from Subcritical Water Extraction on Photodamage in Human Keratinocytes.

The objective of this study was to evaluated the photoprotective effects of porcine placenta extract (PPE) on ultraviolet B (UVB)-induced oxidative stress in human keratinocytes (HaCaT) to evaluate its functional activities as a skin food ingredient. PPE prepared by subcritical water extraction was termed SPE, and subsequently digested by enzymes to prepare E-SPE. Increased intracellular reactive oxygen species (ROS) levels (192.0%) induced by UVB were decreased by SPE and E-SPE. SPE had more effective ROS scavenging activity than E-SPE treatment. UVB treatment increased expression of tissue inhibitor of metalloproteinase 1 (TIMP-1), and this elevated expression was decreased by E-SPE treatment. High-dose treatment with E-SPE (50 and 100 µg/mL) reduced TIMP-1 expression levels of UVB-C (control) to 33.5 and 34.6%, respectively. In contrast, at low SPE doses (1 and 10 µg/mL), the treatment slightly decreased TIMP-1 expression levels to 73.3% and 71.3% of UVB-C, respectively. In conclusion, the present study demonstrated the protective effect of SPE and E-SPE against UVB damage in keratinocytes via ROS scavenging, down-regulating MMP-2 expression and up-regulating TIMP-1 expression. This highlights the potential for SPE as an ingredient in the preparation of functional food against photoaging.

Influence of the antiandrogen flutamide on connexin 43 (Cx43) gene and protein expression in the porcine placenta and uterus during pregnancy.

The study focuses on the expression of connexin 43 (Cx43), a gap junctional protein in the porcine placenta and uterus. The aim was to examine Cx43 mRNA and protein expression after antiandrogen flutamide treatment. Flutamide was injected into pregnant gilts at a daily dose of 50 mg/kg body weight at different stages of pregnancy: between days 43-49 (50 dpc), 83-89 (90 dpc) and 101-107 (108 dpc) of gestation. The animals were sacrificed and tissues were collected one day after the last injection. Cx43 immunostaining was observed in epithelial and stromal cells of the fetal part of the placenta; luminal and glandular epithelial cells of maternal part of the placenta and myometrium of the uterus within placentation sites. Cx43 was also found in glandular epithelium and myometrium of non-placental uterus. Flutamide treatment caused fluctuations in Cx43 expression especially before parturition. Although significant changes in Cx43 mRNA expression were observed only in the fetal part of the placenta, Cx43 protein level was affected within the maternal part of the placenta and non-placental uterus. These results suggest the involvement of androgens in the regulation of Cx43 expression within the feto-maternal compartment in pigs. However, androgen deficiency caused pronounced changes during late pregnancy and before parturition. These results are interesting due to the functional changes in the porcine uterus during the preparturient period that is determined by Cx43 protein.

Anti-fatigue effects of porcine placenta and its amino acids in a behavioral test on mice.

Placenta extracts are used for their health benefits; however, the anti-fatigue effects of placenta have not been elucidated. Thus, we investigated the anti-fatigue effects of porcine placenta extract (PE) and the amino acids present in the PE (glycine, Gly; proline, Pro; glutamic acid, GA; and arginine, Arg) using a forced swimming test (FST) and a tail-suspension test (TST) on mice. Whole PE or individual amino acids decreased immobility times in the FST. PE, Pro, and Arg all lowered blood levels of lactic acid and alanine aminotransferase (ALT). PE and Gly improved glycogen content and catalase activity. As determined from the serum after the FST: PE regulated the effects of interferon (IFN)-γ and tumor necrosis factor (TNF)-α; GA regulated the effects of IFN-γ; Gly and Arg regulated the effects of interleukin (IL)-6; and all of the amino acids present in PE regulated the effects of TNF-α. As determined from the spleen after the FST: Gly and Arg regulated the effects of IL-1β; Gly, Pro, and Arg regulated the effects of IL-6; PE and all of the amino acids present in PE regulated the effects of TNF-α. After the TST, PE and all of the amino acids present in PE reduced immobility duration as well as levels of aspartate aminotransferase and ALT. As determined from the serum after the TST: PE and Gly regulated the effects of TNF-α; Gly and Arg regulated the effects of IL-1β; Gly, Pro, and Arg regulated the effects of IL-6; PE and all of the amino acids present in PE regulated the effects of TNF-α. These results suggest that PE should be considered a candidate anti-fatigue agent.

Altered imprinted gene expression and methylation patterns in mid-gestation aborted cloned porcine fetuses and placentas.

To determine the expression patterns of imprinted genes and their methylation status in aborted cloned porcine fetuses and placentas. RNA and DNA were prepared from fetuses and placentas that were produced by SCNT and controls from artificial insemination. The expression of 18 imprinted genes was determined by quantitative real-time PCR (q-PCR). Bisulfite sequencing PCR (BSP) was conducted to determine the methylation status of PRE-1 short interspersed repetitive element (SINE), satellite DNA and H19 differentially methylated region 3 (DMR3). The weight, imprinted gene expression and genome-wide DNA methylation patterns were compared between the mid-gestation aborted and normal control samples. The results showed hypermethylation of PRE-1 and satellite sequences, the aberrant expression of imprinted genes, and the hypomethylation of H19 DMR3 occurred in mid-gestation aborted fetuses and placentas. Cloned pigs generated by somatic cell nuclear transfer (SCNT) showed a greater ratio of early abortion during mid-gestation than did normal controls because of the incomplete epigenetic reprogramming of the donor cells. Altered expression of imprinted genes and the hypermethylation profile of the repetitive regions (PRE-1 and satellite DNA) may be associated with defective development and early abortion of cloned pigs, emphasizing the importance of epigenetics during pregnancy and implications thereof for patient-specific embryonic stem cells for human therapeutic cloning and improvement of human assisted reproduction.

Immunohistochemical distribution of early pregnancy factor in ovary, oviduct and placenta of pregnant gilts

Early pregnancy factor (EPF) is an immunosuppressant that promotes maternal immune system tolerance of the allogenic fetus. Little is known about localization of this factor in different tissues and nothing has been reported about localization in swine reproductive and placental tissues. We determined the concentration of EPF in serum of gilts and porcine placenta conditioned medium (PPCM). We also analyzed the expression of EPF in different reproductive tissues of pregnant gilts at 10, 30, 60 and 90 days of pregnancy. EPF concentration in serum and PPCM was determined by western blot and densitometry. EPF expression in reproductive tissue was assessed by immunohistochemistry. The highest concentration of EPF was observed at 30 days in serum and PPCM; the concentration was higher in PPCM than in serum at the stages we evaluated. All reproductive tissues from the gestational stages analyzed showed specific labeling of EPF, but this labeling did not appear in non-pregnant gilts. At 30 days pregnancy, the EPF expression in the ovary was predominantly in follicular lutein cells, probably owing to its function as a luteotrophic factor. In the oviduct, EPF was expressed in unciliated secretory epithelial cells and in the cilia of ciliated cells. In the placenta, EPF was expressed in the fetal portion (mesoderm chorioallantois and epithelium of endoderm). EPF acts as an autocrine and paracrine growth factor for the trophoblast during the peri-implantation period.

Optimization of ultrasound-assisted-extraction of porcine placenta water-soluble proteins and evaluation of the antioxidant activity.

Porcine placenta is commonly used in Chinese as a traditional medicine. It has been reported by a number of researchers that the porcine placenta contains many compounds which have good health benefits. Response surface methodology (RSM) was applied to optimize the parameters of ultrasound power, ultrasound treat time, and extraction temperature on the extraction yield of porcine placenta water-soluble proteins (PPWP). The results indicated that, under optimum conditions of ultrasound power 257 w, extraction temperature at 49°C for 7min, the extraction yield of PPWP was 32.7%, which was significantly higher than that of the conventional extraction method (CEM) of 15.0%. The experimental data were fitted to a second-order polynomial equation using multiple regression analysis and the estimated model showed a high capacity of predicting the dependent variables. Although not significantly, the antioxidant activity of PPWP from ultrasound-assist-extraction (UAE) were higher than those from CEM, indicated that UAE had a positive effect or at least no negative effect on the bioactivity of PPWP.

VEGF system expression by immunohistochemistry and real-time RT-PCR study on collared peccary placenta

Vascular endothelial growth factor (VEGF) is known to induce endothelial cell proliferation, to promote cell migration, and to inhibit apoptosis, thus playing a central role in angiogenesis and in the regulation of vasculogenesis. The expression of the VEGF–ligand receptor system was studied in the placenta and uterus of the collared peccary in nonpregnant females in the luteal phase and throughout pregnancy (>35, 75, 115, and 135 days). The material was examined by immunohistochemistry and by real-time reverse transcription polymerase chain reaction. Intense positive immunolabeling was observed for VEGF and its receptors in the uterine epithelium, uterine glands, and trophoblast. The endothelial cells and smooth muscle cells in the maternal and fetal vessels, as well as the connective tissue and mesenchyme, had weak immunoreactivity during all periods of pregnancy. The regression analysis of the real-time polymerase chain reaction results demonstrated cubic behavior, showing a specific time-dependent profile during pregnancy, which increased over the last gestational period to VEGF and VEGFR-1. The relative expression of VEGFR-2 decreased in the middle-third of the pregnancy and increased in late pregnancy. In the collared peccary, the expression of the VEGF–ligand receptor system was similar to that in porcine and ruminant placentas, suggesting that an epitheliochorial placenta has the same physiological and interhemal barrier during vascular gestational development. The expression of VEGF among cells not related to the vascular system, such as those of the uterine epithelium, trophoblast, and uterine glands, suggests a distinct regulatory role for these cells in vasculogenesis and also a different role of VEGF pathway.

Disruption of imprinted gene expression and DNA methylation status in porcine parthenogenetic fetuses and placentas.

Parthenogenetically activated oocytes cannot develop to term in mammals due to the lack of paternal gene expression and failed X chromosome inactivation (XCI). To further characterize porcine parthenogenesis, the expression of 18 imprinted genes was compared between parthenogenetic (PA) and normally fertilized embryos (Con) using quantitative real-time PCR (qRT-PCR). The results revealed that maternally expressed genes were over-expressed, whereas paternally expressed genes were significantly reduced in PA fetuses and placentas. The results of bisulfite sequencing PCR (BSP) demonstrated that PRE-1 and Satellite were hypermethylated in both Con and PA fetuses and placentas, while XIST DMRs were hypomethylated only in PA samples. Taken together, these results suggest that the aberrant methylation profile of XIST DMRs and abnormal imprinted gene expression may be responsible for developmental failure and impaired growth in porcine parthenogenesis.

Promoter characterization and functional association with placenta of porcine MAGEL2.

MAGEL2 (melanoma antigen-like gene 2) is essential for circadian function, metabolism and reproduction in mammals. This study was conducted to investigate transcriptional regulation and functional importance in the placenta of porcine MAGEL2. Quantitative real-time PCR showed that MAGEL2 was highly expressed in porcine hypothalamus, pituitary and placenta (P<0.05). The gene was down-regulated in Meishan but up-regulated in Duroc placentas from 25 days post-coitum (dpc) to 105 dpc (P<0.01). Dual luciferase assay demonstrated that the region -151/+110 had the highest promoter activity. Of the g. -712C>G and g. -708T>C polymorphisms in MAGEL2 promoter, -712C and -708T were observed to be predominant in Large White, Landrace and Duroc populations, while -712G and -708C were predominant in Meishan and Rongchang populations. Moreover, -712C>G and -708T>C had significant effects on MAGEL2 transcription (P<0.05) and placental efficiency (P<0.01). In conclusion, -151/+110 harbors the basal promoter of porcine MAGEL2. The region upstream the basal promoter contains repressive cis-elements. And, MAGEL2 is essential in porcine placenta.

Assessment of fetal cell chimerism in transgenic pig lines generated by Sleeping beauty transposition.

Human cells migrate between mother and fetus during pregnancy and persist in the respective host for long-term after birth. Fetal microchimerism occurs also in twins sharing a common placenta or chorion. Whether microchimerism occurs in multiparous mammals such as the domestic pig, where fetuses have separate placentas and chorions, is not well understood. Here, we assessed cell chimerism in litters of wild-type sows inseminated with semen of transposon transgenic boars. Segregation of three independent monomeric transposons ensured an excess of transgenic over non-transgenic offspring in every litter. Transgenic siblings (n = 35) showed robust ubiquitous expression of the reporter transposon encoding a fluorescent protein, and provided an unique resource to assess a potential cell trafficking to non-transgenic littermates (n = 7) or mothers (n = 4). Sensitive flow cytometry, fluorescence microscopy, and real-time PCR provided no evidence for microchimerism in porcine littermates, or piglets and their mothers in both blood and solid organs. These data indicate that the epitheliochorial structure of the porcine placenta effectively prevents cellular exchange during gestation.

Effects of Concentration and Reaction Time of Trypsin, Pepsin, and Chymotrypsin on the Hydrolysis Efficiency of Porcine Placenta.

This study investigated the effects of three proteases (trypsin, pepsin and chymotrypsin) on the hydrolysis efficiency of porcine placenta and the molecular weight (Mw) distributions of the placental hydrolysates. Because placenta was made up of insoluble collagen, the placenta was gelatinized by applying thermal treatment at 90 ℃ for 1 h and used as the sample. The placental hydrolyzing activities of the enzymes at varying concentrations and incubation times were determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and gel permeation chromatography (GPC). Based on the SDS-PAGE, the best placental hydrolysis efficiency was observed in trypsin treatments where all peptide bands disappeared after 1 h of incubation as compared to 6 h of chymotrypsin. Pepsin hardly hydrolyzed the placenta as compared to the other two enzymes. The Mw distribution revealed that the trypsin produced placental peptides with Mw of 106 and 500 Da. Peptides produced by chymotrypsin exhibited broad ranges of Mw distribution (1-20 kDa), while the pepsin treatment showed Mw greater than 7 kDa. For comparisons of pre-treatments, the subcritical water processing (37.5 MPa and 200 ℃ of raw placenta improved the efficiency of tryptic digestions to a greater level than that of a preheating treatment (90 ℃ for 1 h). Consequently, subcritical water processing followed by enzymatic digestions has the potential of an advanced collagen hydrolysis technique.

Effect of High Pressure on the Porcine Placenral Hydrolyzing Activity of Pepsin, Trypsin and Chymotrypsin.

This study investigated the effects of protease treatments (trypsin, chymotrypsin, and pepsin) under various pressure levels (0.1-300 MPa) for the characteristics of porcine placenta hydrolysates. According to gel electrophoretic patterns, the trypsin showed the best placental hydrolyzing activity followed by chymotrypsin, regardless of the pressure levels. In particular, the peptide bands of tryptic-digested hydrolysate were not shown regardless of applied pressure levels. The peptide bands of hydrolysate treated chymotrypsin showed gradual decreases in molecular weights (Mw) with increasing pressure levels. However, the pepsin did not show any evidences of placental hydrolysis even though the pressure levels were increased to 300 MPa. The gel permeation chromatography (GPC) profiles showed that the trypsin and pepsin had better placental hydrolyzing activities under high pressure (particularly at 200 MPa), with lower Mw distributions of the hydrolysates. Pepsin also tend to lower the Mw of peptides, while the major bands of hydrolysates being treated at 300 MPa were observed at more than 7,000 Da. There were some differences in amino acid compositions of the hydrolysates, nevertheless, the peptides were mainly composed of glycine (Gly), alanine (Ala), hydroxyproline (Hyp) and proline (Pro). Consequently, the results indicate that high pressure could enhance the placental hydrolyzing activities of the selected proteases and the optimum pressure levels at which the maximum protease activity is around 200 MPa.

Elevated expression of vascular endothelial growth factor (VEGF) 120 in parthenogenetic porcine placentas

Most mammalian parthenogenetic embryos are unable to develop to term due to placental defects, potentially caused by decreased vasculogenesis and angiogenesis of the parthenogenetic placenta. Here we have compared the expression status of vascular endothelial growth factor (VEGF) and angiopoietin family members between normally developing and parthenogenetic porcine placentas. The result showed significantly reduced expression of these genes but elevated expression of VEGF 120 in the parthenogenetic porcine placenta (p < 0.05). We postulate that the abnormal expression levels of VEGF and angiopoietin family members and, especially, the elevated expression of VEGF 120 observed in parthenogenetic porcine placentas are related to the early miscarriage of parthenogenetic embryos in pigs.

Shotgun proteomic analysis of porcine colostrum and mature milk

The epitheliochorial nature of the porcine placenta prevents the transfer of maternal immunity. Therefore, ingestion of the colostrum immediately after birth is crucial for neonatal piglets to acquire passive immunity from the sow. We performed a shotgun proteomic analysis of porcine milk to reveal in detail the protein composition of porcine milk. On the basis of the Swiss-Prot database, 113 and 118 proteins were identified in the porcine colostrum and mature milk, respectively, and 50 of these proteins were common to both samples. Some immune-related proteins, including interleukin-18 (IL-18), were unique to the colostrum. The IL-18 concentration in the colostrum and mature milk of four sows was measured to validate the proteomic analysis, and IL-18 was only detected in the colostrum (191.0 ± 53.9 pg/mL) and not in mature milk. In addition, some proteins involved in primary defense, such as azurocidin, which has never been detected in any other mammal's milk, were also identified in the colostrum.

Choriocarcinoma-like tumor in a potbellied pig (Sus scrofa)

A uterine tumor, with histological and immunohistochemical features consistent with those of human choriocarcinoma, was identified in a 10-year-old unmated female pot-bellied pig (Sus scrofa). The tumor showed biphasic proliferation of cytotrophoblast-like cells and syncytiotrophoblast-like cells. Immunohistochemically, the syncytiotrophoblast-like cells were positive for human chorionic gonadotropin, and both types of cells were positive for cytokeratin and negative for vimentin, octamer-binding transcription factor 4, and α-fetoprotein. Because syncytiotrophoblasts are absent in the normal porcine placenta, the tumor was diagnosed as a choriocarcinoma-like tumor.

Flutamide Influences Placental Aldo–Keto Reductase Family 1 Member C1 (AKR1C1) Expression in Pigs

Aldo-keto reductase family 1 member C1 (AKR1C1) catalyses the conversion of progesterone into inactive 20α-dihydroxyprogesterone. It is suggested that AKR1C1 expression in the placenta prevents from the cytotoxic effect of progesterone on foetuses during late pregnancy. The aim of the study was to determine whether the anti-androgen flutamide administered during late pregnancy (83-89days of gestation) or before parturition (101-107days of gestation) influences AKR1C1 expression in the porcine placenta. AKR1C1 mRNA and protein levels were measured using real-time PCR and western blotting, respectively. Immunolocalization of AKR1C1 within placentas was also performed. Flutamide significantly increased AKR1C1 mRNA (p=0.008) and protein (p=0.019) expression only during the pre-parturient period in pigs. AKR1C1 protein was immunolocalized in the epithelial and stromal cells of foetal and maternal part of placenta at both stages of gestation. Following flutamide treatment, the intensity of staining was higher (p=0.045) on day 108 of gestation. In conclusion, porcine placental AKR1C1 expression seems to be regulated by an androgen signalling pathway and may be involved in foetal survival by preventing the detrimental effect of progesterone.

79 UTERINE AND PLACENTAL INTERACTIONS DURING NECROTIC TIP DEVELOPMENT IN THE PIG FROM DAY 22 TO 42 OF GESTATION

Placental development is important for fetal development and nutrient and waste transport. The pig, a litter bearing animal, has an epitheliochorial placenta that forms a noninvasive attachment with the uterine endometrium. Insufficient placental development is one of the primary causes of fetal death and reduced fetal growth after 35 days of gestation. Necrotic tips develop at the distal ends of each allantochorion between Day 22 and 42 of gestation. During this same period, placenta attaches to the uterine endometrium and establish fetal blood supplies and nutrient exchange. The attached placenta is composed of a central highly vascular placental (HVP) region adjacent to the fetus and less vascular placental (LVP) regions on either side to the fetus, the paraplacenta and necrotic tips, which form after 27 days of gestation. The objective of this study was to comprehensively evaluate uterine–placental interactions and necrotic tip development from 22 to 42 days of gestation in the gilt. Gilts (n = 25) were bred by AI at first detection of oestrus (Day 0) and harvested at 22, 27, 32, 37, or 42 days of gestation. Each conceptus was counted and weighed to identify the large, medium, and small fetus in each litter. From these fetuses, HVP and LVP sections of tissue and necrotic tip (no placental attachment) were collected. Intact attached uterus and placenta were collected for histology and preserved in paraformaldehyde. Placentas were then stripped from the endometrium and individually weighed. Hematoxylin and eosin staining of uterine : placental sections was used to identify uterine fold development in the HVP, LVP, and uterine endometrial morphology in the necrotic tip regions. Three folds were measured for depth, width, and area using BQ Nova Prime software (Bioquant Image Analysis, Nashville, TN, USA). Data were analysed using PROC MIXED in SAS (SAS Institute Inc., Cary, NC, USA). Litter size, 12.1 ± 3.4, was similar (P = 0.86) for all days of gestation. Fetal and placental weight increased (P &lt; 0.05) as day of gestation increased. Average fetal weight was similar (P = 0.30) for Day 27 and 32 with a tendency (P = 0.09) to increase by Day 37 before a significant increase at Day 42 (P = 0.002). The placenta increased (P = 0.02) in weight throughout this period of gestation, with the greatest increase in weight between 37 and 42 days of gestation. The LVP zones had no measureable fold formation until Day 32 from most conceptuses, and all LVP zones displayed microfold formation by Day 37. Necrotic tips became apparent after 27 days of gestation. Necrotic tip areas of the uterus had observable modifications from Day 32 to 42 of gestation, developing folds with dramatic changes in endometrial cell size and morphology. This work demonstrated fundamental time points in placental development that correspond to fetal growth and microfold formation. Gestation Day 27 through 32 show limited changes in either fetal growth or increased placental weight; however, significant morphological changes occur throughout the placenta, and even necrotic tips demonstrating the dynamic architecture of the establishing porcine placenta during early gestation.

Effects of Preparation and Drying Methods on the Antioxidant Activity of Enzymatically Hydrolyzed Porcine Placenta Hydrolysates

Porcine placenta was hydrolyzed using papain and trypsin in a combined formulation. An orthogonal experimental design (L9(3)4) was applied in order to obtain porcine placenta hydrolysates (PPH) having the best antioxidant activities. The PPH was dried using vacuum drying (VD), spray drying (SD), and freeze drying (FD) and the effects of these drying methods on the antioxidative activity and solubility were investigated. The DPPH scavenging activity of these PPH reached 44.45 ± 0.46% and the molecular weight of 83.39% of these PPH peptides was below 3000Da. The proportion of highly antioxidative amino acid residues in these PPH reached 60.14%. The VD-PPH showed the lowest antioxidantive activities (DPPH, superoxide and hydroxyl radical scavenging activities, and lecithin liposome antioxidant activity) and the lowest solubility. The FD-PPH exhibited the highest antioxidative properties and the highest solubility. Both the antioxidative properties and the solubility of SD-PPH remained in between those of FD-PPH and VD-PPH. Spray drying can be a method of choice to produce PPH powders due to its higher throughput.

Porcine placenta mitigates protein–energy malnutrition-induced fatigue

ObjectiveFatigue can be caused by a deficiency of nutrition or immune function. The goal of this study was to identify the effects of porcine placenta extract (PPE) and its constituents, amino acids (glutamic acid, glycine, arginine, and proline), on protein–energy malnutrition (PEM)-induced fatigue. MethodsMice were administered a PEM diet and came to immunodeficient status. Simultaneously, the mice were administered PPE or amino acids and a forced swimming test (FST) was performed. We analyzed the levels of fatigue-related factors in serum, splenocytes, and muscles. ResultsIn the FST, PPE or amino acids significantly decreased immobility times compared with the PEM diet. PPE or amino acids also significantly decreased the serum levels of fatigue-related factors after the FST. Additionally, PPE significantly decreased the levels of fatigue-related muscle parameters after the FST. In this in vitro study, PPE increased the mRNA and protein expression of Ki-67 and promoted the proliferation of splenocytes. PPE or amino acids significantly increased the levels of intracellular calcium and the translocation into the nucleus of nuclear factor of activated T-cells cytoplasmic in stimulated splenocytes. PPE or amino acids significantly decreased the production of fatigue-related inflammatory cytokines in the stimulated splenocytes. Additionally, the translocated levels of nuclear factor-κB in the nucleus and the degradation of the inhibitory protein, IκBα, in the cytosol were inhibited by PPE or amino acids. ConclusionThese results demonstrate that PPE and its constituents regulate PEM-induced fatigue through improving levels of immunity and decreasing fatigue-related factors. PPE may be a potential agent for a recovery from fatigue.