OBJECTIVE: To investigate the development and dynamic changes of cysts in the brain of mice following infection with different forms of Toxoplasma gondii, so as to provide insights into for toxoplasmosis prevention and control.
METHODS: ICR mice at ages of 6 to 8 weeks, each weighing 20 to 25 g, were intraperitoneally injected with tachyzoites of the T. gondii PRU strain at a dose of 1 × 105 tachyzoites per mouse, orally administered with cysts at a dose of 20 oocysts per mouse or oocysts at a dose of 200 oocysts per mouse for modeling chronic T. gondii infection in mice, and the clinical symptoms and survival of mice were observed post-infection. Mice were orally infected with T. gondii cysts at doses of 10 (low-dose group), 20 (medium-dose group), 40 cysts per mouse (high-dose group), and the effect of different doses of T. gondii infections on the number of cysts was examined in the mouse brain. Mice were orally administered with T. gondii cysts at a dose of 20 cysts per mouse, and grouped according to gender (female and male) and time points of infections (20, 30, 60, 90, 120, 150, 180 days post-infection), and the effects of gender and time points of infections on the number of cysts was examined in the mouse brain. In addition, mice were divided into the tachyzoite group (Group T), the first-generation cyst group (Group C1), the second-generation cyst group (Group C2), the third-generation cyst (Group C3) and the fourth-generation cyst group (Group C4). Mice in the Group T were intraperitoneally injected with T. gondii tachyzoites at a dose of 1 × 105 tachyzoites per mouse, and the cysts were collected from the mouse brain tissues 30 days post-infection, while mice in the Group C1 were orally infected with the collected cysts at a dose of 30 cysts per mouse. Continuous passage was performed by oral administration with cysts produced by the previous generation in mice, and the effect of continuous passage on the number of cysts was examined in the mouse brain.
RESULTS: Following infection with T. gondii tachyzoites, cysts and oocysts in mice, obvious clinical symptoms were observed on days 6 to 13 and mice frequently died on days 7 to 12. The survival rates of mice were 67.0%, 87.0% and 53.0%, and the mean numbers of cysts were (516.0 ± 257.2), (1 203.0 ± 502.0) and (581.0 ± 183.1) in the mouse brain (F = 11.94, P < 0.01) on day 30 post-infection with T. gondii tachyzoites, cysts and oocysts, respectively, and the numbers of cysts in the brain tissues were significantly lower in mice infected with T. gondii tachyzoites and oocysts than in those infected with cysts (all P values < 0.01). The survival rates of mice were 87.0%, 87.0% and 60.0%, and the mean numbers of cysts were (953.0 ± 355.5), (1 084.0 ± 474.3) and (1 113.0 ± 546.0) in the mouse brain in the low-, medium- and high-dose groups on day 30 post-infection, respectively (F = 0.42, P > 0.05). The survival rates of male and female mice were 73.0% and 80.0%, and the mean numbers of cysts were (946.4 ± 411.4) and (932.1 ± 322.4) in the brain tissues of male and female mice, respectively (F = 1.63, P > 0.05). Following continuous passage, the mean numbers of cysts were (516.0 ± 257.2), (1 203.0 ± 502.0), (896.8 ± 332.3), (782.5 ± 423.9) and (829.2 ± 306.0) in the brain tissues of mice in the T, C1, C2, C3 and C4 groups, respectively (F = 4.82, P < 0.01), and the number of cysts was higher in the mouse brain in Group 1 than in Group T (P < 0.01). Following oral administration of 20 T. gondii cysts in mice, cysts were found in the moues brain for the first time on day 20 post-infection, and the number of cysts gradually increased over time, peaked on days 30 and 90 post-infection and then gradually decreased; however, the cysts were still found in the mouse brain on day 180 post-infection.
CONCLUSIONS: There is a higher possibility of developing chronic T. gondii infection in mice following infection with cysts than with oocysts or tachyzoites and the most severe chronic infection is seen following infection with cysts. The number of cysts does not correlate with the severity of chronic T. gondii infection, and the number of cysts peaks in the mouse brain on days 30 and 90 post-infection.
［摘要］ 目的 观察不同形态刚地弓形虫感染后小鼠脑内包囊形成及其动态变化, 为弓形虫病防控提供依据。方法 取 6~8周龄ICR小鼠 (20~25 g) 建立慢性弓形虫感染模型, 其中弓形虫PRU株速殖子按1 × 105个/只剂量腹腔注射感染小鼠, 包囊和卵囊分别按20、200个/只剂量通过灌胃针口服感染小鼠, 观察感染后小鼠临床症状和存活情况。分别以10 (低剂 量组) 、20 (中剂量组) 、40个包囊/只 (高剂量组) 剂量感染小鼠, 观察弓形虫不同感染剂量对小鼠脑内包囊数量的影响。 将小鼠按性别 (雌、雄性) 、感染时间 (感染后20、30、60、90、120、150、180 d) 分组, 按20个/只剂量口服弓形虫包囊后, 分别 观察性别和感染时间对小鼠脑内包囊数量的影响。将小鼠分成速殖子组 (T组) 、包囊1代组 (C1组) 、包囊2代组 (C2 组) 、包囊3代组 (C3组) 、包囊4代组 (C4组); T组小鼠按1 × 105个/只剂量腹腔注射弓形虫速殖子, 感染后第30天处死小 鼠并收集其脑组织内包囊, 再按20个/只感染C1组小鼠。此后每一代小鼠均采用上一代所产生包囊进行口服连续传代, 观察连续传代对小鼠脑内弓形虫包囊数量的影响。结果 以弓形虫速殖子、包囊、卵囊分别感染小鼠, 感染第6~13天 小鼠出现明显临床症状、感染第 7~12 天小鼠出现集中死亡。感染第 30 天时, 感染速殖子、包囊、卵囊的小鼠存活率分 别为67.0%、87.0%、53.0%, 平均脑内包囊数量分别为 (516.0 ± 257.2) 、 (1 203.0 ± 502.0) 、 (581.0 ± 183.1) 个, 差异有统计 学意义 (F = 11.94, P < 0.01), 感染速殖子、卵囊的小鼠脑内包囊数低于感染包囊的小鼠 (P 均< 0.01) 。感染后第30天, 低、中、高剂量组小鼠存活率分别为87.0%、87.0%、60.0%, 平均脑内包囊数量分别为 (953.0 ± 355.5) 、 (1 084.0 ± 474.3) 、 (1 113.0 ± 546.0) 个, 差异无统计学意义 (F = 0.42, P > 0.05); 雄、雌性组小鼠存活率分别为73.0%和80.0%, 平均脑内包 囊数量分别为 (946.4 ± 411.4) 、 (932.1 ± 322.4) 个, 差异无统计学意义 (F = 1.63, P > 0.05) 。通过连续传代感染后, T、C1、 C2、C3、C4组小鼠平均脑内包囊数量分别为 (516.0 ± 257.2) 、 (1 203.0 ± 502.0) 、 (896.8 ± 332.3) 、 (782.5 ± 423.9) 、 (829.2 ± 306.0) 个, 差异有统计学意义 (F = 4.82, P < 0.01); C1组小鼠脑内包囊数高于速殖子组, 差异有统计学意义 (P < 0.01) 。 小鼠口服20个包囊后, 感染第20天首次查见脑内弓形虫包囊, 随感染时间延长脑内包囊数量逐渐增加; 至感染第30、90 天时, 脑内包囊数量分别达峰值, 此后逐步下降, 至感染第180天时仍能查见脑内包囊。结论 刚地弓形虫包囊较速殖 子、卵囊感染后形成慢性感染的可能性更高, 且慢性感染程度亦最严重; 感染弓形虫包囊数量与慢性感染严重程度无关; 脑内包囊形成数量于感染第30天和90天时达高峰。.

In this study, we examined the potential antidepressant-like effects of Chinese quince fruit extract (Chaenomeles sinensis fruit extract, CSFE) in an in vivo model induced by repeated injection of corticosterone (CORT)-induced depression. HPLC analysis determined that chlorogenic acid (CGA), neo-chlorogenic acid (neo-CGA), and rutin (RT) compounds were major constituents in CSFE. Male ICR mice (5 weeks old) were orally administered various doses (30, 100, and 300 mg/kg) of CSFE and selegiline (10 mg/kg), a monoamine oxidase B (MAO-B) inhibitor, as a positive control following daily intraperitoneal injections of CORT (40 mg/kg) for 21 days. In our results, mice treated with CSFE exhibited significant improvements in depressive-like behaviors induced by CORT. This was evidenced by reduced immobility times in the tail suspension test and forced swim test, as well as increased step-through latency times in the passive avoidance test. Indeed, mice treated with CSFE also exhibited a significant decrease in anxiety-like behaviors as measured by the elevated plus maze test. Moreover, molecular docking analysis indicated that CGA and neo-CGA from CSFE had stronger binding to the active site of MAO-B. Our results indicate that CSFE has potential antidepressant effects in a mouse model of repeated injections of CORT-induced depression.

Astatine (211At) is a cyclotron-produced alpha emitter with a physical half-life of 7.2 h. In our previous study, the 211At-labeled prostate-specific membrane antigen (PSMA) compound ([211At]PSMA-5) exhibited excellent tumor growth suppression in a xenograft model. We conducted preclinical biodistribution and toxicity studies for the first-in-human clinical trial. [211At]PSMA-5 was administered to both normal male ICR mice (n = 85) and cynomolgus monkeys (n = 2). The mice were divided into four groups for the toxicity study: 5 MBq/kg, 12 MBq/kg, 35 MBq/kg, and vehicle control, with follow-ups at 1 day (n = 10 per group) and 14 days (n = 5 per group). Monkeys were observed 24 h post-administration of [211At]PSMA-5 (9 MBq/kg). Blood tests and histopathological examinations were performed at the end of the observation period. Blood tests in mice indicated no significant myelosuppression or renal dysfunction. However, the monkeys displayed mild leukopenia 24 h post-administration. Despite the high accumulation in the kidneys and thyroid, histological analysis revealed no abnormalities. On day 1, dose-dependent single-cell necrosis/apoptosis was observed in the salivary glands of mice and intestinal tracts of both mice and monkeys. Additionally, tingible body macrophages in the spleen and lymph nodes indicated phagocytosis of apoptotic B lymphocytes. Cortical lymphopenia (2/10) in the thymus and a decrease in the bone marrow cells (9/10) were observed in the 35 MBq/kg group in mice. These changes were transient, with no irreversible toxicity observed in mice 14 days post-administration. This study identified no severe toxicities associated with [211At]PSMA-5, highlighting its potential as a next-generation targeted alpha therapy for prostate cancer. The sustainable production of 211At using a cyclotron supports its applicability for clinical use.

BACKGROUND: At present, neonatal hypoxic-ischemic encephalopathy (HIE), especially moderate to severe HIE, is a challenging disease for neonatologists to treat, and new alternative/complementary treatments are urgently needed. The neuroinflammatory cascade triggered by hypoxia-ischemia (HI) insult is one of the core pathological mechanisms of HIE. Early inhibition of neuroinflammation provides long-term neuroprotection. Plant-derived monomers have impressive anti-inflammatory effects. Aloesin (ALO) has been shown to have significant anti-inflammatory and antioxidant effects in diseases such as ulcerative colitis, but its role in HIE is unclear. To this end, we conducted a series of experiments to explore the potential mechanism of ALO in preventing and treating brain damage caused by HI insult.
METHODS: Hypoxic-ischemic brain damage (HIBD) was induced in 7-day-old Institute of Cancer Research (ICR) mice, which were then treated with 20 mg/kg ALO. The neuroprotective effects of ALO on HIBD and the underlying mechanism were evaluated through neurobehavioral testing, infarct size measurement, apoptosis detection, protein and messenger RNA level determination, immunofluorescence, and molecular docking.
RESULTS: ALO alleviated the long-term neurobehavioral deficits caused by HI insult; reduced the extent of cerebral infarction; inhibited cell apoptosis; decreased the levels of the inflammatory factors interleukin (IL)-1β, IL-6, and tumor necrosis factor-α; activated microglia and astrocytes; and downregulated the protein expression of members in the TLR4 signaling pathway. In addition, molecular docking showed that ALO can bind stably to TLR4.
CONCLUSIONS: ALO ameliorated HIBD in neonatal mice by inhibiting the neuroinflammatory response mediated by TLR4 signaling.

OBJECTIVE: To assess the effects of repeated mild traumatic brain injury (rmTBI) in the parietal cortex on neuronal morphology and synaptic plasticity in the medulla oblongata of mice.
METHODS: Thirty-two male ICR mice were randomly divided into sham operation group (n=8) and rmTBI group (n=24). The mice in the latter group were subjected to repeated mild impact injury of the parietal cortex by a free-falling object. The mice surviving the injuries were evaluated for neurological deficits using neurological severity scores (NSS), righting reflex test and forced swimming test, and pathological changes of the neuronal cells in the medulla oblongata were observed with HE and Nissl staining. Western blotting and immunofluorescence staining were used to detect the expressions of neuroligin 1(NLG-1) and postsynaptic density protein 95(PSD-95) in the medulla oblongata of the mice that either survived rmTBI or not.
RESULTS: None of the mice in the sham-operated group died, while the mortality rate was 41.67% in rmTBI group. The mice surviving rmTBI showed significantly reduced NSS, delayed recovery of righting reflex, increased immobility time in forced swimming test (P < 0.05), and loss of Nissl bodies; swelling and necrosis were observed in a large number of neurons in the medulla oblongata, where the expression levels of NLG-1 and PSD-95 were significantly downregulated (P < 0.05). The mice that did not survive rmTBI showed distorted and swelling nerve fibers and decreased density of neurons in the medulla oblongina with lowered expression levels of NLG-1 and PSD-95 compared with the mice surviving the injuries (P < 0.01).
CONCLUSIONS: The structural and functional anomalies of the synapses in the medulla oblongata may contribute to death and neurological impairment following rmTBI in mice.

Vibrio vulnificus causes life-threatening wound and gastrointestinal infections, mediated primarily by the production of a Multifunctional-Autoprocessing Repeats-In-Toxin (MARTX) toxin. The most commonly present MARTX effector domain, the Makes Caterpillars Floppy-like (MCF) toxin, is a cysteine protease stimulated by host adenosine diphosphate (ADP) ribosylation factors (ARFs) to autoprocess. Here, we show processed MCF then binds and cleaves host Ras-related proteins in brain (Rab) guanosine triphosphatases within their C-terminal tails resulting in Rab degradation. We demonstrate MCF binds Rabs at the same interface occupied by ARFs. Moreover, we show MCF preferentially binds to ARF1 prior to autoprocessing and is active to cleave Rabs only subsequent to autoprocessing. We then use structure prediction algorithms to demonstrate that structural composition, rather than sequence, determines Rab target specificity. We further determine a crystal structure of aMCF as a swapped dimer, revealing an alternative conformation we suggest represents the open, activated state of MCF with reorganized active site residues. The cleavage of Rabs results in Rab1B dispersal within cells and loss of Rab1B density in the intestinal tissue of infected mice. Collectively, our work describes an extracellular bacterial mechanism whereby MCF is activated by ARFs and subsequently induces the degradation of another small host guanosine triphosphatase (GTPase), Rabs, to drive organelle damage, cell death, and promote pathogenesis of these rapidly fatal infections.

We investigated whether calorie restriction (CR) enhances metabolic adaptations to endurance training (ET). Ten-week-old male Institute of Cancer Research (ICR) mice were fed ad libitum or subjected to 30% CR. The mice were subdivided into sedentary and ET groups. The ET group performed treadmill running (20-25 m/min, 30 min, 5 days/week) for 5 weeks. We found that CR decreased glycolytic enzyme activity and monocarboxylate transporter (MCT) 4 protein content, while enhancing glucose transporter 4 protein content in the plantaris and soleus muscles. Although ET and CR individually increased citrate synthase activity in the plantaris muscle, the ET-induced increase in respiratory chain complex I protein content was counteracted by CR. In the soleus muscle, mitochondrial enzyme activity and protein levels were increased by ET, but decreased by CR. It has been suggested that CR partially interferes with skeletal muscle adaptation to ET.

Atmospheric particulate matter (PM) exacerbates the risk factor for Alzheimer\'s and Parkinson\'s diseases (PD) by promoting the alpha-synuclein (α-syn) pathology in the brain. However, the molecular mechanisms of astrocytes involvement in α-syn pathology underlying the process remain unclear. This study investigated PM with particle size <200 nm (PM0.2) exposure-induced α-syn pathology in ICR mice and primary astrocytes, then assessed the effects of mammalian target of rapamycin inhibitor (PP242) in vitro studies. We observed the α-syn pathology in the brains of exposed mice. Meanwhile, PM0.2-exposed mice also exhibited the activation of glial cell and the inhibition of autophagy. In vitro study, PM0.2 (3, 10 and 30 µg/mL) induced inflammatory response and the disorders of α-syn degradation in primary astrocytes, and lysosomal-associated membrane protein 2 (LAMP2)-mediated autophagy underlies α-syn pathology. The abnormal function of autophagy-lysosome was specifically manifested as the expression of microtubule-associated protein light chain 3 (LC3II), cathepsin B (CTSB) and lysosomal abundance increased first and then decreased, which might both be a compensatory mechanism to toxic α-syn accumulation induced by PM0.2. Moreover, with the transcription factor EB (TFEB) subcellular localization and the increase in LC3II, LAMP2, CTSB, and cathepsin D proteins were identified, leading to the restoration of the degradation of α-syn after the intervention of PP242. Our results identified that PM0.2 exposure could promote the α-syn pathological dysregulation in astrocytes, providing mechanistic insights into how PM0.2 increases the risk of developing PD and highlighting TFEB/LAMP2 as a promising therapeutic target for antagonizing PM0.2 toxicity.

The safety of the carrageenan (CGN) consumption as a food additive is under debate, with negative effects being associated with the products of hydrolysis of CGN. Moreover, there is an increasing need to integrate gut microbiome analysis in the scientific risk assessment of food additives. The objective of this study was to test the effects of CGN consumption on the gut microbiota and the intestinal homeostasis of young male and female mice. Female and male ICR-CD1 mice (8 weeks old) orally received 540 mg kg-1 day-1 of CGN, representing the maximum-level exposure assessment scenario surveyed for children, over the course of two weeks. Fecal material and peritoneal immune cells were analyzed to determine changes in the fecal microbiota, based on the analysis of bacterial 16S rRNA gene amplicon sequences and short-chain fatty acid (SCFA) concentrations, and some immune functions and redox parameters of peritoneal leukocytes. Non-significant microbiota taxonomical changes associated with CGN intake were found in the mouse stools, resulting the housing time in an increase in bacterial groups belonging to the Bacteroidota phylum. The PICRUSt2 functional predictions showed an overall increase in functional clusters of orthologous genes (COGs) involved in carbohydrate transport and metabolism. A significant increase in the cytotoxicity of fecal supernatants was observed in CGN-fed mice, which correlated with worsening of immune functions and oxidative parameters. The altered immunity and oxidative stress observed in young mice after the consumption of CGN, along with the fecal cytotoxicity shown towards intestinal epithelial cells, may be associated with the gut microbiota\'s capacity to degrade CGN. The characterization of the gut microbiota\'s ability to hydrolyze CGN should be included in the risk assessment of this food additive.

OBJECTIVE: Aspartame (L-aspartyl L-phenylalanine methyl ester) is an artificial sweetener widely used as a sugar substitute. There are concerns regarding the effects of high aspartame doses on the kidney owing to oxidative stress; however, whether the maximum allowed dose of aspartame in humans affects the kidneys remains unknown. Therefore, in this study, we investigated whether the maximum allowed dose of aspartame in humans affects the kidneys.
METHODS: In this study, animals were fed a folate-deficient diet to mimic human aspartame metabolism. Eight-week-old ICR mice were divided into control (CTL), 40 mg/kg/day of aspartame-administered (ASP), folate-deficient diet (FD), and 40 mg/kg/day of aspartame-administered with a folate-deficient diet (FD + ASP) groups. Aspartame was administered orally for eight weeks. Thereafter, we evaluated aspartame\'s effect on kidneys via histological analysis.
RESULTS: There were no differences in serum creatinine and blood urea nitrogen levels between the CTL and ASP groups or between the FD and FD + ASP groups. There was no histological change in the kidneys in any group. The expression of superoxide dismutase and 4-hydroxy-2-nonenal in the kidney did not differ between the CTL and ASP groups or the FD and FD + ASP groups.
CONCLUSIONS: Our findings indicate that the allowed doses of aspartame in humans may not affect kidney function or oxidative states.