PSD-95 in CA1 Area Regulates Spatial Choice Depending on Age

Cognitive processes that require spatial information rely on synaptic plasticity in the dorsal CA1 area (dCA1) of the hippocampus. Since the function of the hippocampus is impaired in aged individuals, it remains unknown how aged animals make spatial choices. Here, we used IntelliCage to study behavioral processes that support spatial choices of aged female mice living in a group. As a proxy of training-induced synaptic plasticity, we analyzed the morphology of dendritic spines and the expression of a synaptic scaffold protein, PSD-95. We observed that spatial choice training in young adult mice induced correlated shrinkage of dendritic spines and downregulation of PSD-95 in dCA1. Moreover, long-term depletion of PSD-95 by shRNA in dCA1 limited correct choices to a reward corner, while reward preference was intact. In contrast, old mice used behavioral strategies characterized by an increased tendency for perseverative visits and social interactions. This strategy resulted in a robust preference for the reward corner during the spatial choice task. Moreover, training decreased the correlation between PSD-95 expression and the size of dendritic spines. Furthermore, PSD-95 depletion did not impair place choice or reward preference in old mice. Thus, our data indicate that while young mice require PSD-95-dependent synaptic plasticity in dCA1 to make correct spatial choices, old animals observe cage mates and stick to a preferred corner to seek the reward. This strategy is resistant to the depletion of PSD-95 in the CA1 area. Overall, our study demonstrates that aged mice combine alternative behavioral and molecular strategies to approach and consume rewards in a complex environment.SIGNIFICANCE STATEMENT It remains poorly understood how aging affects behavioral and molecular processes that support cognitive functions. It is, however, essential to understand these processes to develop therapeutic interventions that support successful cognitive aging. Our data indicate that while young mice require PSD-95-dependent synaptic plasticity in dCA1 to make correct spatial choices (i.e., choices that require spatial information), old animals observe cage mates and stick to a preferred corner to seek the reward. This strategy is resistant to the depletion of PSD-95 in the CA1 area. Overall, our study demonstrates that aged mice combine alternative behavioral and molecular strategies to approach and consume rewards in a complex environment. Second, the contribution of PSD-95-dependent synaptic functions in spatial choice changes with age.     

Heterozygous germline BLM mutations increase susceptibility to asbestos and mesothelioma

Rare biallelic BLM gene mutations cause Bloom syndrome. Whether BLM heterozygous germline mutations (BLM^+/−) cause human cancer remains unclear. We sequenced the germline DNA of 155 mesothelioma patients (33 familial and 122 sporadic). We found 2 deleterious germline BLM^+/− mutations within 2 of 33 families with multiple cases of mesothelioma, one from Turkey (c.569_570del; p.R191Kfs*4) and one from the United States (c.968A>G; p.K323R). Some of the relatives who inherited these mutations developed mesothelioma, while none with nonmutated BLM were affected. Furthermore, among 122 patients with sporadic mesothelioma treated at the US National Cancer Institute, 5 carried pathogenic germline BLM^+/− mutations. Therefore, 7 of 155 apparently unrelated mesothelioma patients carried BLM^+/− mutations, significantly higher (P = 6.7E-10) than the expected frequency in a general, unrelated population from the gnomAD database, and 2 of 7 carried the same missense pathogenic mutation c.968A>G (P = 0.0017 given a 0.00039 allele frequency). Experiments in primary mesothelial cells from Blm^+/− mice and in primary human mesothelial cells in which we silenced BLM revealed that reduced BLM levels promote genomic instability while protecting from cell death and promoted TNF-α release. Blm^+/− mice injected intraperitoneally with asbestos had higher levels of proinflammatory M1 macrophages and of TNF-α, IL-1β, IL-3, IL-10, and IL-12 in the peritoneal lavage, findings linked to asbestos carcinogenesis. Blm^+/− mice exposed to asbestos had a significantly shorter survival and higher incidence of mesothelioma compared to controls. We propose that germline BLM^+/− mutations increase the susceptibility to asbestos carcinogenesis, enhancing the risk of developing mesothelioma.

In the United States, the incidence rate of mesothelioma varies between fewer than one case per 100,000 persons in states with no asbestos industry to two to three cases per 100,000 persons in states with an asbestos industry (1, 2). Asbestos causes DNA damage and apoptosis (3) and promotes a chronic inflammatory reaction that supports the emergence of malignant cells (4). Fortunately, only a small fraction of exposed individuals develop mesothelioma; for example, 4.6% of deaths in miners who worked in asbestos mines for over 10 y were caused by mesothelioma (1). Therefore, multiple cases of mesothelioma in the same family are rare and suggest genetic predisposition (5). In 2001, we discovered that susceptibility to mesothelioma was transmitted in a Mendelian fashion across multiple generations in some Turkish families exposed to the carcinogenic fiber erionite, pointing to gene × environment interaction (G×E) as the cause (6). In 2011, we discovered that carriers of heterozygous germline BRCA1-associated protein–1 (BAP1) mutations (BAP1^+/−) developed mesothelioma and uveal melanoma (5), findings expanded and confirmed by us and by multiple research teams (reviewed in refs. 1, 7, 8). Moreover, heterozygous germline Bap1 mutations (Bap1^+/−) significantly increased susceptibility to asbestos-induced mesothelioma in mice (9, 10), evidence of G×E. Reduced BAP1 levels impair DNA repair (11) as well as different forms of cell death (3, 12) and induce metabolic alterations (13–15) that together favor cancer development and growth.Recent studies revealed that mesothelioma may also develop among carriers of germline mutations of additional tumor-suppressor genes that cause well-defined cancer syndromes, including MLH1 and MLH3 (Lynch syndrome), TP53 (Li–Fraumeni syndrome), and BRCA1-2 (Breast and Ovarian Cancer syndrome) (16, 17). When all germline mutations are combined, it has been estimated that about 12% of mesotheliomas occur in carriers of heterozygous germline mutations of BAP1, the most frequent mutation among patients with mesothelioma, or of other tumor suppressors. Some of these mutations may sensitize the host to asbestos carcinogenesis, according to a G×E scenario (17). Thus, presently, mesothelioma is considered an ideal model to study G×E in cancer (17). As part of the Healthy Nevada Project (HNP), we are studying G×E in northern Nevada, a region with an unusually high risk of exposure to carcinogenic minerals and arsenic, which may be related to the high cancer rates in this region (18). We are investigating genetic variants that may increase cancer risk upon exposure to carcinogens to implement preventive strategies.Biallelic mutations of the Bloom syndrome gene (BLM) cause Bloom syndrome, an autosomal-recessive tumor predisposition syndrome characterized by pre- and postnatal growth deficiency, photosensitivity, type 2 diabetes, and greatly increased risk of developing various types of cancers. BLM is a RecQ helicase enzyme that modulates DNA replication and repair of DNA damage by homologous recombination (19). In patients affected by Bloom syndrome, the absence of the BLM protein causes chromosomal instability, increased number of sister chromatid exchanges, and increased numbers of micronuclei (20–22). In addition, BLM is required for p53-mediated apoptosis (23), a process critical to eliminate cells that have accumulated DNA damage. Impaired DNA repair together with altered apoptosis resulted in increased cancer incidence (17, 24). Of course, inactivating germline BLM heterozygous (BLM^+/−) mutations are much more common than biallelic BLM (BLM^−/−) mutations, with an estimated frequency in the general population of 1 in 900 based on data from the Exome Aggregation Consortium (25). BLM^+/− mutation carriers do not show an obvious phenotype; however, some studies have suggested that carriers of these mutations may have an increased cancer risk (17, 24). Mice carrying Blm^+/− mutations are prone to develop a higher rate of malignancies in the presence of contributing factors, such as concurrent heterozygous mutations of the adenomatous polyposis coli (Apc) gene, or upon infection with murine leukemia virus (26). However, in studies in which Blm^+/− mice were crossed with tuberous sclerosis 1-deficient (Tsc1^+/−) mice that are predisposed to renal cystadenomas and carcinomas, Wilson et al. found that Tsc1^+/− Blm^+/− mice did not show significantly more renal cell carcinomas compared with Tsc1^+/− Blm^WT mice (27). In humans, a large study involving 1,244 patients with colon cancer and 1,839 controls of Ashkenazi Jewish ancestry, in which BLM^+/− frequency is as high as 1 in 100 individuals (28), suggested that carriers of germline BLM^+/− mutations might have a twofold increase in colorectal cancer (CRC) (29). A smaller study did not confirm these results, but reported a trend of increasing incidence of adenomas—premalignant lesions—among BLM^+/− mutation carriers (30). In addition, BLM^+/− mutations were found overrepresented among early-onset (<45 y old) CRC patients (25). Other studies associated BLM^+/− mutations to an increased risk of breast (31, 32) and prostate cancer (33), but the low power of these studies hampered definite conclusions. In summary, it appears possible that BLM^+/− mutations may increase cancer risk in the presence of contributing factors.     

Arachidyl amido cholanoic acid improves liver glucose and lipid homeostasis in nonalcoholic steatohepatitis via AMPK and mTOR regulation

BACKGROUND Arachidyl amido cholanoic acid(Aramchol) is a potent downregulator of hepatic stearoyl-CoA desaturase 1(SCD1) protein expression that reduces liver triglycerides and fibrosis in animal models of steatohepatitis. In a phase IIb clinical trial in patients with nonalcoholic steatohepatitis(NASH), 52 wk of treatment with Aramchol reduced blood levels of glycated hemoglobin A1c, an indicator of glycemic control.AIM To assess lipid and glucose metabolism in mouse hepatocytes and in a NASH mouse model [induced with a 0.1% methionine and choline deficient diet(0.1 MCD)] after treatment with Aramchol.METHODS Isolated primary mouse hepatocytes were incubated with 20 μmol/L Aramchol or vehicle for 48 h. Subsequently, analyses were performed including Western blot, proteomics by mass spectrometry, and fluxomic analysis with ~(13)C-uniformly labeled glucose. For the in vivo part of the study, male C57 BL/6 J mice were randomly fed a control or 0.1 MCD for 4 wk and received 1 or 5 mg/kg/d Aramchol or vehicle by intragastric gavage for the last 2 wk. Liver metabolomics were assessed using ultra-high-performance liquid chromatography-time of flight-MS for the determination of glucose metabolism-related metabolites.RESULTS Combination of proteomics and Western blot analyses showed increased AMPK activity while the activity of nutrient sensor mTORC1 was decreased by Aramchol in hepatocytes. This translated into changes in the content of their downstream targets including proteins involved in fatty acid(FA) synthesis and oxidation [PACCα/β(S79), SCD1, CPT1A/B, HADHA, and HADHB], oxidative phosphorylation(NDUFA9, NDUFB11, NDUFS1, NDUFV1, ETFDH, and UQCRC2), tricarboxylic acid(TCA) cycle(MDH2, SUCLA2, and SUCLG2), and ribosome(P-p70S6K[T389] and P-S6[S235/S236]). Flux experiments with ~(13)C uniformely labeled glucose showed that TCA cycle cataplerosis was reduced by Aramchol in hepatocytes, as indicated by the increase in the number of rounds that malate remained in the TCA cycle. Finally, liver metabolomic analysis showed that glucose homeostasis was improved by Aramchol in 0.1 MCD fed mice in a dose-dependent manner, showing normalization of glucose, G6P, F6P, UDP-glucose, and Rbl5 P/Xyl5 P.CONCLUSION Aramchol exerts its effect on glucose and lipid metabolism in NASH through activation of AMPK and inhibition of mTORC1, which in turn activate FA β-oxidation and oxidative phosphorylation.