Regulatory effects of Trichinella spiralis serpin-type serine protease inhibitor on endoplasmic reticulum stress and oxidative stress in host intestinal epithelial cells.

Endoplasmic reticulum stress (ERS) and oxidative stress (OS) are adaptive responses of the body to stressor stimulation. Although it has been verified that Trichinella spiralis (T. spiralis) can induce ERS and OS in the host, their association is still unclear. Therefore, this study explored whether T. spiralis-secreted serpin-type serine protease inhibitor (TsAdSPI) is involved in regulating the relationship between ERS and OS in the host intestine. In this study, mice jejunum and porcine small intestinal epithelial cells (IECs) were detected using qPCR, western blotting, immunohistochemistry (IHC), immunofluorescence (IF), and detection kits. The results showed that ERS- and OS-related indexes changed significantly after TsAdSPI stimulation, and Bip was located in IECs, indicating that TsAdSPI could induce ERS and OS in IECs. After the use of an ERS inhibitor, OS-related indexes were inhibited, suggesting that TsAdSPI-induced OS depends on ERS. When the three ERS signalling pathways, ATF6, IRE1, and PERK, were sequentially suppressed, OS was only regulated by the PERK pathway, and the PERK-eif2α-CHOP-ERO1α axis played a key role. Similarly, the expression of ERS-related indexes and the level of intracellular Ca2+ were inhibited after adding the OS inhibitor, and the expression of ERS-related indexes decreased significantly after inhibiting calcium transfer. This finding indicated that TsAdSPI-induced OS could affect ERS by promoting Ca2+ efflux from the endoplasmic reticulum. The detection of the ERS and OS sequences revealed that OS occurred before ERS. Finally, changes in apoptosis-related indexes were detected, and the results indicated that TsAdSPI-induced ERS and OS could regulate IEC apoptosis. In conclusion, TsAdSPI induced OS after entering IECs, OS promoted ERS by enhancing Ca2+ efflux, and ERS subsequently strengthened OS by activating the PERK-eif2α-CHOP-ERO1α axis. ERS and OS induced by TsAdSPI synergistically promoted IEC apoptosis. This study provides a foundation for exploring the invasion mechanism of T. spiralis and the pathogenesis of host intestinal dysfunction after invasion.

Monitoring of Chlorophyll Content of Potato in Northern Shaanxi Based on Different Spectral Parameters.

Leaf chlorophyll content (LCC) is an important physiological index to evaluate the photosynthetic capacity and growth health of crops. In this investigation, the focus was placed on the chlorophyll content per unit of leaf area (LCCA) and the chlorophyll content per unit of fresh weight (LCCW) during the tuber formation phase of potatoes in Northern Shaanxi. Ground-based hyperspectral data were acquired for this purpose to formulate the vegetation index. The correlation coefficient method was used to obtain the "trilateral" parameters with the best correlation between potato LCCA and LCCW, empirical vegetation index, any two-band vegetation index constructed after 0-2 fractional differential transformation (step size 0.5), and the parameters with the highest correlation among the three spectral parameters, which were divided into four combinations as model inputs. The prediction models of potato LCCA and LCCW were constructed using the support vector machine (SVM), random forest (RF) and back propagation neural network (BPNN) algorithms. The results showed that, compared with the "trilateral" parameter and the empirical vegetation index, the spectral index constructed by the hyperspectral reflectance after differential transformation had a stronger correlation with potato LCCA and LCCW. Compared with no treatment, the correlation between spectral index and potato LCC and the prediction accuracy of the model showed a trend of decreasing after initial growth with the increase in differential order. The highest correlation index after 0-2 order differential treatment is DI, and the maximum correlation coefficients are 0.787, 0.798, 0.792, 0.788 and 0.756, respectively. The maximum value of the spectral index correlation coefficient after each order differential treatment corresponds to the red edge or near-infrared band. A comprehensive comparison shows that in the LCCA and LCCW estimation models, the RF model has the highest accuracy when combination 3 is used as the input variable. Therefore, it is more recommended to use the LCCA to estimate the chlorophyll content of crop leaves in the agricultural practices of the potato industry. The results of this study can enhance the scientific understanding and accurate simulation of potato canopy spectral information, provide a theoretical basis for the remote sensing inversion of crop growth, and promote the development of modern precision agriculture.

Effects of Trichinella spiralis and its serine protease inhibitors on autophagy of host small intestinal cells.

In eukaryotes, autophagy is induced as an innate defense mechanism against pathogenic microorganisms by self-degradation. Although trichinellosis is a foodborne zoonotic disease, there are few reports on the interplay between Trichinella spiralissurvival strategies and autophagy-mediated host defense. Therefore, this study focused on the association between T. spiralis and autophagy of host small intestinal cells. In this study, the autophagy-related indexes of host small intestinal cells after T. spiralis infection were detected using transmission electron microscopy, hematoxylin and eosin staining, immunohistochemistry, quantitative real-time polymerase chain reaction, and Western blotting. The results showed that autophagosomes and autolysosomes were formed in small intestinal cells, intestinal villi appeared edema, epithelial compactness was decreased, microtubule-associated protein 1A/1B-light chain 3B (LC3B) was expressed in lamina propria stromal cells of small intestine, and the expression of autophagy-related genes and proteins was changed significantly, indicating that T. spiralis induced autophagy of host small intestinal cells. Then, the effect of T. spiralis on autophagy-related pathways was explored by Western blotting. The results showed that the expression of autophagy-related pathway proteins was changed, indicating that T. spiralis regulated autophagy by affecting autophagy-related pathways. Finally, the roles of T. spiralis serine protease inhibitors (TsSPIs), such as T. spiralis Kazal-type SPI (TsKaSPI) and T. spiralis Serpin-type SPI (TsAdSPI), were further discussed in vitro and in vivo experiments. The results revealed that TsSPIs induced autophagy by influencing autophagy-related pathways, and TsAdSPI has more advantages. Overall, our results indicated that T. spiralis induced autophagy of host small intestinal cells, and its TsSPIs play an important role in enhancing autophagy flux by affecting autophagy-related pathways. These findings lay a foundation for further exploring the pathogenesis of intestinal dysfunction of host after T. spiralis infection, and also provide some experimental and theoretical basis for the prevention and treatment of trichinellosis.

Exosome-delivered miR-153 from Trichinella spiralis promotes apoptosis of intestinal epithelial cells by downregulating Bcl2.

Trichinellosis, a helminthic zoonosis, exhibits a cosmopolitan distribution and is a public health concern. In previous studies, it was reported that the exosomes secreted by Trichinella spiralis larvae (TsExos) largely affected cell biological activities. miRNAs, as exosome-delivered cargoes, affect the biological activities of the host by targeting genes. The present study aimed to elucidate the mechanisms by which miRNAs interact with intestinal epithelial cells. First, a miRNA library of TsExos was constructed; then, based on high-throughput miRNA sequencing results, miR-153 and its predicted target genes, namely, Agap2, Bcl2 and Pten, were selected for follow-up studies. The dual-luciferase reporter assays revealed that miR-153 directly targeted Bcl2 and Pten. Furthermore, real-time qPCR and Western blotting revealed that only Bcl2 was downregulated by TsExo-delivered miR-153 in porcine intestinal epithelial cells (IPEC-J2). Bcl2, an important antiapoptotic protein, plays an essential role in cell apoptosis as a common intersecting molecule of various signal transduction pathways. Therefore, we hypothesized that miR-153 derived from TsExos causes cell apoptosis by targeting Bcl2. The results suggested that miR-153 could induce apoptosis, reduce mitochondrial membrane potential, affect cell proliferation, and cause damage and substantial oxidative stress. Furthermore, miR-153 coincubated with IPEC-J2 cells stimulated the accumulation of the proapoptotic proteins Bax and Bad, which belong to the Bcl2 family of proteins, and the apoptosis-implementing proteins Caspase 9 and Caspase 3. Moreover, studies have suggested that miR-153 can promote apoptosis by regulating the MAPK and p53 signalling pathways involved in apoptosis. Thus, exosome-mediated miR-153 delivery secreted by T. spiralis could induce apoptosis and affect the MAPK and p53 signalling pathways by downregulating Bcl2 in IPEC-J2 cells. The study highlights the mechanisms underlying the invasion of T. spiralis larva.

Intervention effects of Trichinella spiralis excretory-secretory antigens on allergic asthma in mice.

Allergic asthma is a chronic, heterogeneous and inflammatory respiratory disease, and there are few medicines at present. An increasing number of studies indicate that Trichinella spiralis (T. spiralis) and its excretory-secretory (ES) antigens are inflammatory modulator. Therefore, this study focused on the effects of T. spiralis ES antigens on allergic asthma. Asthma model was established by sensitizing mice with ovalbumin antigen (OVA) and aluminum hydroxide (Al[OH]3), the asthmatic mice were interfered using T. spiralis 43 kDa protein (Ts43), T. spiralis 49 kDa protein (Ts49), and T. spiralis 53 kDa protein (Ts53), the important components of ES antigens, to establish ES antigens intervention models. Then, asthma symptom changes, weight changes, and lung inflammation of mice were evaluated. The results showed that ES antigens could relieve symptoms, weight loss, and lung inflammation caused by asthma in the mice, and the effect of combined intervention of Ts43, Ts49, and Ts53 was better. Finally, the effects of ES antigens on type 1 helper T (Th1) and type 2 helper T (Th2) immune responses, and the differentiation direction of T lymphocytes in mice were discussed by detecting Th1 and Th2 cell-related factors and the ratio of CD4+/CD8+ T cells. The results suggested that the ratio of CD4+/CD8+ T cells decreased and the ratio of Th1/Th2 cells increased. In conclusion, this study indicated that T. spiralis ES antigens could mitigate allergic asthma in the mice by changing the differentiation direction of CD4+ and CD8+ T cells and regulating the imbalance of Th1/Th2 cells ratio.

Individual and combined effects of heat and drought and subsequent recovery on winter wheat (Triticum aestivum L.) photosynthesis, nitrogen metabolism, cell osmoregulation, and yield formation.

Extreme temperatures and droughts are considered as the two main factors that limit wheat growth and production. Although responses of wheat plants to heat and drought stress have been extensively investigated, little is known about the extent to which wheat plants can recover after stress relief. In this study, a winter wheat pot experiment was conducted to evaluate the growth, physiological activities, and yield formation responses of wheat to stress and recovery periods under heat stress (36 °C, daily maximum temperature), drought (45-55% of soil water holding capacity), and combined stress conditions. Heat and drought stress significantly reduced photosynthesis, leaf relative water content (LRWC), leaf water potential (LWPnoon), and nitrogen metabolism enzyme activities and increased electrolyte leakage. These parameters showed significant interactions between heat and drought stress. Beneficial osmoregulation of membrane stability was observed in stressed plants because of the accumulation of proline and soluble sugars. Within a range of stresses, the abovementioned physiological processes of individual heat- and drought-stressed plants recovered to levels comparable to those of the control. The recovery capacities of the physiological traits decreased gradually with increasing stress duration, particularly under combined stress. The recovery of LWPnoon and LRWC contributed to the improved photosynthetic performance after stress relief. The combined stress caused greater yield losses than individual heat and drought stress, which was mainly attributed to low levels of thousand grain weight (TGW), the number of grains per ear, and the grain filling rate. After stress relief, the recovery of proline content, glutamine synthetase activity, photosynthetic rate, and LRWC were closely associated with grain yield and thousand grain weight. Collectively, these findings contribute to a better understanding of the coordinated responses of winter wheat during the combined heat and drought stress and recovery periods.

Photosynthetic, antioxidant activities, and osmoregulatory responses in winter wheat differ during the stress and recovery periods under heat, drought, and combined stress.

There will be longer and more intense periods of heat and drought stress in the future for terrestrial ecosystems. Although the responses of wheat plants to heat and drought stress alone have been extensively investigated, little is known about the extent to which their recovery can be assured after stress relief. In this study, a winter wheat pot experiment was conducted to investigate the changes in photosynthetic performance, antioxidant activity, osmoregulation, and membrane lipid peroxidation under heat stress (36 °C), drought (45-55% of soil water holding capacity), and combined stress conditions. The results showed that heat and drought stress significantly reduced the photosynthetic rate and the contents of chlorophyll and carotenoid. Superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and glutathione reductase (GR) activities were greatly activated by heat and drought stress to scavenge overproduced superoxide anion (O2-). Plants exhibited positive osmoregulation through the synthesis of soluble protein (SP), soluble sugar (SS), and proline (Pro) to improve membrane stability. Within a range of stress, combined heat and drought stress exhibited significant interactive effects in the above mentioned indicators. After stress relief, the majority of physiological processes were reversible, as indicated by the effective recovery of pigment contents, photosynthetic rate, antioxidant enzyme activities, osmoregulatory substance contents, and O2- production. Antioxidant enzyme activities tended to increase after recovering from 12 days of combined stress, whereas they were still not effective in mitigating oxidative damage. High levels of O2- and malondialdehyde (MDA) and a low relative growth rate during the recovery confirmed the irreversible damage caused by combined heat and drought stress. ROC (receiver operating characteristic) analysis indicated that GR and SS could accurately detect individual heat and drought stress that wheat plants were suffering or had suffered (AUC = 0.812-0.965), while POD and Pro had greater potential for diagnosing combined heat and drought stress (AUC = 0.871-0.958). Physiological indicators of stress tolerance were closely related to the photosynthetic rate during the stress, particularly Pro and GR. Collectively, the physiological processes of plants are reversible within a certain range of stress. POD, GR, Pro, and SS play vital roles in identifying and resisting heat, drought, and combined stress, and the recovery of these indicators contributed to improving photosynthesis and thereby increasing wheat growth. Our research contributes to the understanding of the underlying physiological mechanisms of plants in response to combined heat and drought stress and after stress relief.

Effects of exosomes derived from Trichinella spiralis infective larvae on intestinal epithelial barrier function.

Muscle larvae of Trichinella spiralis parasitize the host intestinal epithelium. The mechanisms of exosomes participating in the invasion of T. spiralis muscle larvae are unclear. Hence, the purpose of this study was to explore the effect of exosomes derived from T. spiralis infective larvae (TsExos) on the barrier function of porcine small intestinal epithelial cells (IPEC-J2). First, TsExos were successfully obtained, and their ingestion by epithelial cells was validated. Furthermore, the optimal induction condition was determined by the CCK8 kit, and we found that exposure to 150 µg/mL TsExos for 12/24 h decreased the viability of IPEC-J2 cells by 30%. Based on this outcome, the effects of TsExos on cell biological processes and tight junctions were studied. After coincubation of TsExos and IPEC-J2 cells, the results showed a significant increase in the content of FITC-dextran and in the levels of lactate dehydrogenase (LDH) and reactive oxygen species (ROS). The rate of apoptosis increased by 12.57%, and nuclear pyknosis and nuclear rupture were observed. After the cells were induced by TsExos, the expression of IL-1 was upregulated, but the expression of IL-10, TGF-ß, TLR-5, MUC-1 and MUC-2 was downregulated. TsExo induction also led to a decrease in the levels of ZO-1, CLDN-3, and OCLN. In conclusion, TsExos are involved in several cellular biological processes, and they function by disrupting physiological and biochemical processes, hyperactivating innate immunity, and damaging tight junctions.

Regulatory effects of Trichinella spiralis and a serine protease inhibitor on the endoplasmic reticulum stress response of intestinal epithelial cells.

The accumulation of unfolded or misfolded proteins in the endoplasmic reticulum can cause an endoplasmic reticulum stress (ERS) response. If ERS continues or cannot be alleviated, it will cause the production of proapoptotic factors and eventually lead to apoptosis. Therefore, this study mainly explored whether Trichinella spiralis Kazal-type serine protease inhibitor (TsKaSPI) contributed to the invasion of intestinal epithelial cells during the infectious stage of T. spiralis by regulating ERS. First, in the T. spiralis infection model, H&E staining was used to analyse the damage to jejunum tissue, a TUNEL assay was used to examine cell apoptosis, and the expression of ERS-related and apoptosis-related molecules was also measured. The results showed that ERS occurred during the intestinal phase of T. spiralis infection, while remission began during the cyclic phase. Then, we selected TsKaSPI, one of the important components of T. spiralis ES antigens, for in vitro experiments. The results showed that TsKaSPI could induce apoptosis in a porcine small intestinal epithelial cell line (IPEC cells) by activating ERS and promote activation of the NF-κB signalling pathway. Inhibition experiments confirmed that the occurrence of ERS was accompanied by the activation of NF-κB, and the two processes regulated each other. Finally, we conducted in vivo experiments and administered TsKaSPI to mice. The results confirmed that TsKaSPI could activate ERS and lead to apoptosis in intestinal epithelial cells. In conclusion, T. spiralis infection and TsKaSPI can promote cell apoptosis by activating the ERS response in intestinal epithelial cells and activate the NF-κB signalling pathway to promote the occurrence and development of inflammation.

[Astragalus protects the testis following testicular torsion/detorsion in rats].

OBJECTIVE: To investigate the protective effect of astragalus on testis tissues following unilateral testicular torsion/detorsion. METHODS: Thirty healthy adult Wistar rats were equally randomized into Group A (sham-operation control), B (torsion/detorsion) and C (torsion/detorsion plus intraperitoneal injection of astragalus). The testicular torsion/detorsion model was established by the Turner method. All the rats were fed under the same condition for 7 days and sacrificed, and the torsional testes were harvested for the detection of germ cell apoptosis, glutathione activity and the level of malonic diethylaldehyde (MDA). RESULTS: The apoptosis indexes (AI) of spermatogenic cells in the torsional testes were (5.82 +/- 1.21), (36.18 +/- 8.40) and (20.39 +/- 3.57) in Group A, B and C, significantly higher in Group B and C than in A (P < 0.05) and in Group B than in C (P < 0.05). Significant differences were observed in glutathione activity in the ipsilateral testes among Group A (48.03 +/- 2.01), B (30.93 +/- 1.25), C (38.44 +/- 1.06) U/mg (P< 0.05), as well as in the level of MDA, (1.43 +/- 0.17), (3.98 +/- 0.36), (2.57 +/- 0.53) nmol/ml, among the three groups (P < 0.05). CONCLUSION: Astragalus could significantly reduce the apoptosis of spermatogenic cells, decrease the level of lipid peroxidation and protect glutathione activity in the torsional testis.