Neuroprotective Effects of Casein-Derived Peptide Met-Lys-Pro (MKP) in a Hypertensive Model.

We have previously reported that casein hydrolysate, CH-3, from bovine milk and casein-derived tripeptide Met-Lys-Pro (MKP) has ACE inhibitory activity and reduces blood pressure. In this study, we investigated the therapeutic effects of MKP in a hypertensive rat model (7-week-old male SHRSP/Izm rats). For long term evaluation, rats were fed either a diet containing CH-3 or normal diet. The survival rate of SHRSP rats was significantly improved by intake of CH-3 for 181 days. For short term evaluation, rats were orally administered synthetic tripeptide MKP or distilled water for 4 weeks. MRI study demonstrated that hemorrhagic lesions were observed in two of five rats in the control group, while no hemorrhagic lesions were observed in the MKP group. Volumetric analysis using MRI revealed that MKP administration inhibited atrophy of diencephalic regions. Histological examinations revealed that hemorrhage areas and astrogliosis in the hippocampus and cerebral cortex were lower in the MKP group than in the control group. Gene expression analysis indicated that MKP administration reduced expression of genes related to cerebral circulation insufficiency such as immune responses (Cd74 and Prkcd), response to hypoxia (Ddit4, Apold1, and Prkcd), reactive oxygen species metabolic process (Ddit4 and Pdk4), and apoptotic process (Ddit4, Prkcd, and Sgk1), suggesting that MKP administration prevented cerebral ischemia associated with hypertension. In addition, some genes encoding responses to hormone stimulus (Fos, Dusp1, and Sik1) were also downregulated. Serum aldosterone and corticosterone levels were also significantly decreased following MKP administration. The present study indicates that MKP shows neuroprotective effects in SHRSP rats by regulating cerebral circulation insufficiency and corticoid levels. MKP administration may therefore be a potential therapeutic strategy for hypertensive brain diseases such as cerebrovascular disease

Pharmaceutical Potential of Casein-Derived Tripeptide Met-Lys-Pro: Improvement in Cognitive Impairments and Suppression of Inflammation in APP/PS1 Mice

Background:Tripeptide Met-Lys-Pro (MKP), a component of casein hydrolysates, has effective angiotensin-converting enzyme (ACE) inhibitory activity. Brain angiotensin II enzyme activates the NADPH oxidase complex via angiotensin II receptor type 1 (AT1) and enhances oxidative stress injury. ACE inhibitors improved cognitive function in Alzheimer\u27s disease (AD) mouse models and previous clinical trials. Thus, although undetermined, MKP may be effective against pathological amyloid-β (Aβ) accumulation-induced cognitive impairment.Objective:The current study aimed to investigate the potential of MKP as a pharmaceutical against AD by examining MKP\u27s effect on cognitive function and molecular changes in the brain using double transgenic (APP/PS1) mice.Methods:Experimental procedures were conducted in APP/PS1 mice (n = 38) with a C57BL/6 background. A novel object recognition test was used to evaluate recognition memory. ELISA was used to measure insoluble Aβ40, Aβ42, and TNF-α levels in brain tissue. Immunohistochemical analysis allowed the assessment of glial cell activation in MKP-treated APP/PS1 mice.Results:The novel object recognition test revealed that MKP-treated APP/PS1 mice showed significant improvement in recognition memory. ELISA of brain tissue showed that MKP significantly reduced insoluble Aβ40, Aβ42, and TNF-α levels. Immunohistochemical analysis indicated the suppression of the marker for microglia and reactive astrocytes in MKP-treated APP/PS1 mice.Conclusion:Based on these results, we consider that MKP could ameliorate pathological Aβ accumulation-induced cognitive impairment in APP/PS1 mice. Furthermore, our findings suggest that MKP potentially contributes to preventing cognitive decline in AD

Administration of Non-Absorbable Antibiotics to Pregnant Mice to Perturb the Maternal Gut Microbiota Is Associated with Alterations in Offspring Behavior

<div><p>There is increasing evidence that the gut microbiota plays a major role in host health and disease. In this study, we examined whether perturbation of the maternal gut microbiota during pregnancy, induced by administration of non-absorbable antibiotics to pregnant dams, influences the behavior of offspring. Terminal restriction fragment length polymorphism analyses of fecal bacterial composition showed that the relative abundance of the bacterial order <i>Lactobacillales</i> was lower in offspring born from antibiotic-treated dams (20.7±3.4%) than in control offspring (42.1±6.2%) at P24, while the relative abundance of the bacterial family <i>Clostridium</i> subcluster XIVa was higher in offspring born from antibiotic-treated dams (34.2±5.0%) than in control offspring (16.4±3.3%). Offspring born from antibiotic-treated dams exhibited low locomotor activity in both familiar and novel environments, and preferred to explore in the peripheral area of an unfamiliar field at postnatal week 4. At postnatal weeks 7–8, no difference was observed in the level of locomotor activity between control offspring and offspring from antibiotic-treated dams, while the tendency for the offspring from antibiotic-treated dams to be less engaged in exploring the inside area was still observed. The behavioral phenotypes of the offspring from antibiotic-treated dams at postnatal week 4 could be rescued to a considerable extent through fostering of these offspring by normal dams from postnatal day 1. Although the detailed underlying mechanisms are not fully elucidated, the present results suggest that administration of non-absorbable antibiotics to pregnant dams to perturb the maternal gut microbiota during pregnancy leads to alterations in the behavior of their offspring.</p></div

Behaviors of AB offspring at PW4.

<p>(<b>a</b>) Data for 24-h cycle activity in a familiar environment for control offspring and AB offspring at P28–P32. Throughout the dark phase (20:00 to 8:00), the mean activity of AB offspring is generally lower than that of control offspring. Significant differences are detected for 01:00–01:59 (*¹P = 0.043, Mann–Whitney test) and 04:00–04:59 (*²P = 0.0362, Welch’s corrected <i>t</i>-test). Control, n = 29 from 8 dams; AB, n = 18 from 5 dams. (<b>b</b>) Schematic diagram showing the areas in the open field. In the entire field, which is a square of 48 cm per side, two nested squares (7.8-cm square and 24.6-cm square) are assumed. Area A (white square in the center of the field), area B (black area), and area C are then defined so that the three areas do not overlap with one another. (<b>c</b>) Locomotor activity during 2h in the open field test of control offspring and AB offspring at P28–P32. AB offspring show less spontaneous locomotor activity in the entire area of the open field than control offspring throughout the test. **¹P = 0.0041, ***²P = 0.0001 (<i>t</i>-test), **³P = 0.0016 (Welch’s corrected <i>t</i>-test); control, n = 22 from 7 dams; AB, n = 18 from 5 dams. (<b>d</b>) Locomotor activity in the inside area (area A+B) of the field in the open field test. **¹P = 0.0045 (Welch’s corrected <i>t</i>-test), **²P = 0.0011, ****³P<0.0001, **⁴P = 0.0012 (Mann–Whitney test). (<b>e</b>) Locomotor activity in the innermost area (area A) of the field. *¹P = 0.0356, ***²P = 0.0004, ***³P = 0.0009 (Mann–Whitney test). AB offspring prefer to explore the peripheral area of the field compared with control offspring. (<b>f</b>) Number of times of rearing, considered to be an exploratory behavior evoked by novel stimuli, observed during 2 h in the open field test. **P = 0.0023 (Welch’s corrected <i>t</i>-test).</p