Axonal transport of Frizzled5 by Alcadein α-containing vesicles is associated with kinesin-1.

Alcadein α (Alcα) and amyloid-ß protein precursor (APP) are cargo receptors that associate vesicles with kinesin-1. These vesicles, which contain either Alcα or APP, transport various proteins/cargo molecules into axon nerve terminals. Here, we analyzed immune-isolated Alcα- and APP-containing vesicles of adult mouse brains with LC-MS/MS and identified proteins present in vesicles that contained either Alcα or APP. Among these proteins, Frizzled-5 (Fzd5), a Wnt receptor, was detected mainly in Alcα vesicles. Although colocalization ratios of Fzd5 with Alcα are low in the neurites of differentiating neurons by a low expression of Fzd5 in embryonic brains, the suppression of Alcα expression decreased the localization of Fzd5 in neurites of primary cultured neurons. Furthermore, Fzd5-EGFP expressed in primary cultured neurons was preferentially transported in axons with the transport velocities of Alcα vesicles. In synaptosomal fractions of adult-mice brains that express higher levels of Fzd5, the amount of Fzd5 and the phosphorylation level of calcium/calmodulin-dependent protein kinase-II were reduced in the Alcα-deficient mice. These results suggest that reduced transport of Fzd5 by Alcα-containing vesicles associated with kinesin-1 in axon terminals may impair the response to Wnt ligands in the noncanonical Ca2+-dependent signal transduction pathway at nerve terminals of mature neurons.

Brain p3-Alcß peptide restores neuronal viability impaired by Alzheimer's amyloid ß-peptide.

We propose a new therapeutic strategy for Alzheimer's disease (AD). Brain peptide p3-Alcß37 is generated from the neuronal protein alcadein ß through cleavage of γ-secretase, similar to the generation of amyloid ß (Aß) derived from Aß-protein precursor/APP. Neurotoxicity by Aß oligomers (Aßo) is the prime cause prior to the loss of brain function in AD. We found that p3-Alcß37 and its shorter peptide p3-Alcß9-19 enhanced the mitochondrial activity of neurons and protected neurons against Aßo-induced toxicity. This is due to the suppression of the Aßo-mediated excessive Ca2+ influx into neurons by p3-Alcß. Successful transfer of p3-Alcß9-19 into the brain following peripheral administration improved the mitochondrial viability in the brain of AD mice model, in which the mitochondrial activity is attenuated by increasing the neurotoxic human Aß42 burden, as revealed through brain PET imaging to monitor mitochondrial function. Because mitochondrial dysfunction is common in the brain of AD patients alongside increased Aß and reduced p3-Alcß37 levels, the administration of p3-Alcß9-19 may be a promising treatment for restoring, protecting, and promoting brain functions in patients with AD.

Accumulation of amyloid-ß in the brain of mouse models of Alzheimer's disease is modified by altered gene expression in the presence of human apoE isoforms during aging.

Apolipoprotein E4 (apoE4) is a risk factor for Alzheimer's disease (AD). Here, we investigated brain amyloid-ß (Aß) accumulation throughout the aging process in an amyloid precursor protein (APP) knock-in (KI) mouse model of AD that expresses human APPNL-G-F with or without human apoE4 or apoE3. Brain Aß42 levels were significantly lower in 9-month-old mice that express human isoforms of apoE than in age-matched APP-KI control mice. Linear accumulation of Aß42 began in 5-month-old apoE4 mice, and a strong increase in Aß42 levels was observed in 21-month-old apoE3 mice. Aß42 levels in cerebroventricular fluid were higher in apoE3 than in apoE4 mice at 6-7 months of age, suggesting that apoE3 is more efficient at clearing Aß42 than apoE4 at these ages. However, apoE3 protein levels were lower than apoE4 protein levels in the brains of 21-month-old apoE3 and apoE4 mice, respectively, which may explain the rapid increase in brain Aß42 burden in apoE3 mice. We identified genes that were downregulated in a human apoE-dependent (apoE4 > apoE3) and age-dependent (apoE3 = apoE4) manner, which may regulate brain Aß burden and/or AD progression. Analysis of gene expression in AD mouse models helps identify molecular mechanisms of pleiotropy by the human APOE gene during aging.

γ-Secretase structure and activity are modified by alterations in its membrane localization and ambient environment.

γ-Secretase cleaves type I transmembrane proteins in a hydrophobic membrane environment following ectodomain shedding. Mutations in PSEN genes, encoding the catalytic subunits of γ-secretase, presenilins, are the most common cause of familial Alzheimer's disease (ad). Pathogenic mutations in PSEN genes increase production of longer and neurotoxic amyloid-ß (Aß) by intramembrane cleavage of membrane-associated amyloid-ß protein precursor (APP) carboxyl-terminal fragment ß, which is generated via primary cleavage of APP by ß-site APP cleaving enzyme 1. The longer Aß is prone to aggregate and accumulate in the brain; however, the accumulation of Aß in brain is also a pathological feature of sporadic ad. Increased pathogenic Aß generation, even in the absence of pathogenic PSEN gene mutations, is one of proposed mechanisms for sporadic ad pathogenesis. γ-Secretase digests substrates in the transmembrane region, generating Aß peptide intermediates of various lengths. The end products, shorter Aß40 and Aß38 peptides, are less neurotoxic, whereas PSEN gene mutations increase the production ratio of longer, neurotoxic Aß species such as Aß42, an intermediate in Aß38 production. γ-Secretase activity or structures is altered because of its aberrant membrane localization or changes in the ambient environment such as luminal acidification. Interestingly, γ-secretase has a pH sensor in presenilins.

Suppression of amyloid-ß secretion from neurons by cis-9, trans-11-octadecadienoic acid, an isomer of conjugated linoleic acid.

Two common conjugated linoleic acids (LAs), cis-9, trans-11 CLA (c9,t11 CLA) and trans-10, cis-12 CLA (t10,c12 CLA), exert various biological activities. However, the effect of CLA on the generation of neurotoxic amyloid-ß (Aß) protein remains unclear. We found that c9,t11 CLA significantly suppressed the generation of Aß in mouse neurons. CLA treatment did not affect the level of ß-site APP-cleaving enzyme 1 (BACE1), a component of active γ-secretase complex presenilin 1 amino-terminal fragment, or Aß protein precursor (APP) in cultured neurons. BACE1 and γ-secretase activities were not directly affected by c9,t11 CLA. Localization of BACE1 and APP in early endosomes increased in neurons treated with c9,t11 CLA; concomitantly, the localization of both proteins was reduced in late endosomes, the predominant site of APP cleavage by BACE1. The level of CLA-containing phosphatidylcholine (CLA-PC) increased dramatically in neurons incubated with CLA. Incorporation of phospholipids containing c9,t11 CLA, but not t10,c12 CLA, into the membrane may affect the localization of some membrane-associated proteins in intracellular membrane compartments. Thus, in neurons treated with c9,t11 CLA, reduced colocalization of APP with BACE1 in late endosomes may decrease APP cleavage by BACE1 and subsequent Aß generation. Our findings suggest that the accumulation of c9,t11 CLA-PC/LPC in neuronal membranes suppresses the production of neurotoxic Aß in neurons.

Extracellular Release of ILEI/FAM3C and Amyloid-ß Is Associated with the Activation of Distinct Synapse Subpopulations.

BACKGROUND: Brain amyloid-ß (Aß) peptide is released into the interstitial fluid (ISF) in a neuronal activity-dependent manner, and Aß deposition in Alzheimer's disease (AD) is linked to baseline neuronal activity. Although the intrinsic mechanism for Aß generation remains to be elucidated, interleukin-like epithelial-mesenchymal transition inducer (ILEI) is a candidate for an endogenous Aß suppressor. OBJECTIVE: This study aimed to access the mechanism underlying ILEI secretion and its effect on Aß production in the brain. METHODS: ILEI and Aß levels in the cerebral cortex were monitored using a newly developed ILEI-specific ELISA and in vivo microdialysis in mutant human Aß precursor protein-knockin mice. ILEI levels in autopsied brains and cerebrospinal fluid (CSF) were measured using ELISA. RESULTS: Extracellular release of ILEI and Aß was dependent on neuronal activation and specifically on tetanus toxin-sensitive exocytosis of synaptic vesicles. However, simultaneous monitoring of extracellular ILEI and Aß revealed that a spontaneous fluctuation of ILEI levels appeared to inversely mirror that of Aß levels. Selective activation and inhibition of synaptic receptors differentially altered these levels. The evoked activation of AMPA-type receptors resulted in opposing changes to ILEI and Aß levels. Brain ILEI levels were selectively decreased in AD. CSF ILEI concentration correlated with that of Aß and were reduced in AD and mild cognitive impairment. CONCLUSION: ILEI and Aß are released from distinct subpopulations of synaptic terminals in an activity-dependent manner, and ILEI negatively regulates Aß production in specific synapse types. CSF ILEI might represent a surrogate marker for the accumulation of brain Aß.

Amyloidogenic processing of amyloid ß protein precursor (APP) is enhanced in the brains of alcadein α-deficient mice.

Alzheimer's disease (AD) is a very common neurodegenerative disorder, chiefly caused by increased production of neurotoxic ß-amyloid (Aß) peptide generated from proteolytic cleavage of ß-amyloid protein precursor (APP). Except for familial AD arising from mutations in the APP and presenilin (PSEN) genes, the molecular mechanisms regulating the amyloidogenic processing of APP are largely unclear. Alcadein α/calsyntenin1 (ALCα/CLSTN1) is a neuronal type I transmembrane protein that forms a complex with APP, mediated by the neuronal adaptor protein X11-like (X11L or MINT2). Formation of the ALCα-X11L-APP tripartite complex suppresses Aß generation in vitro, and X11L-deficient mice exhibit enhanced amyloidogenic processing of endogenous APP. However, the role of ALCα in APP metabolism in vivo remains unclear. Here, by generating ALCα-deficient mice and using immunohistochemistry, immunoblotting, and co-immunoprecipitation analyses, we verified the role of ALCα in the suppression of amyloidogenic processing of endogenous APP in vivo We observed that ALCα deficiency attenuates the association of X11L with APP, significantly enhances amyloidogenic ß-site cleavage of APP, especially in endosomes, and increases the generation of endogenous Aß in the brain. Furthermore, we noted amyloid plaque formation in the brains of human APP-transgenic mice in an ALCα-deficient background. These results unveil a potential role of ALCα in protecting cerebral neurons from Aß-dependent pathogenicity in AD.

γ-Secretase Activity Is Associated with Braak Senile Plaque Stages.

Amyloid ß-proteins (Aßs) Aß1-42 and Aß1-43 are converted via two product lines of γ-secretase to Aß1-38 and Aß1-40. This parallel stepwise processing model of γ-secretase predicts that Aß1-42 and Aß1-43, and Aß1-38 and Aß1-40 are proportional to each other, respectively. To obtain further insight into the mechanisms of parenchymal Aß deposition, these four Aß species were quantified in insoluble fractions of human brains (Brodmann areas 9 to 11) at various Braak senile plaque (SP) stages, using specific enzyme-linked immunosorbent assays. With advancing SP stages, the amounts of deposited Aß1-43 in the brain increased proportionally to those of Aß1-42. Similarly, the amounts of deposited Aß1-38 correlated with those of Aß1-40. Surprisingly, the ratios of deposited Aß1-38/Aß1-42 and Aß1-40/Aß1-43 were proportional and discriminated the Braak SP stages accurately. This result indicates that the generation of Aß1-38 and Aß1-40 decreased and the generation of Aß1-42 and Aß1-43 increased with advancing SP stages. Thus, Aßs deposition might depend on γ-secretase activity, as it does in the cerebrospinal fluid. Here, the extracted γ-secretase from Alzheimer disease brains generates an amount of Aß1-42 and Aß1-43 compared with cognitively normal brains. This refractory γ-secretase localized in detergent-solubilized fractions from brain cortices. But activity modulated γ-secretase, which decreases Aß1-42 and Aß1-43 in the cerebrospinal fluid, localized in detergent-insoluble fractions. These drastic alterations reflect Aß situation in Alzheimer disease brains.

Enhanced amyloid-ß generation by γ-secretase complex in DRM microdomains with reduced cholesterol levels.

A neuropathologic hallmark of Alzheimer's disease (AD) is the presence of senile plaques that contain neurotoxic amyloid-ß protein (Aß) species, which are generated by the cleavage of amyloid ß-protein precursor by secretases such as the γ-secretase complex, preferentially located in detergent-resistant membrane (DRM) regions and comprising endoproteolysed amino- and carboxy-terminal fragments of presenilin, nicastrin, anterior pharynx defective 1 and presenilin enhancer 2. Whereas some of familial AD patients harbor causative PSEN mutations that lead to more generation of neurotoxic Aß42, the contribution of Aß generation to sporadic/late-onset AD remains unclear. We found that the carboxy-terminal fragment of presenilin 1 was redistributed from DRM regions to detergent-soluble membrane (non-DRM) regions in brain tissue samples from individuals with sporadic AD. DRM fractions from AD brain sample had the ability to generate significantly more Aß and had a lower cholesterol content than DRM fractions from non-demented control subjects. We further demonstrated that lowering the cholesterol content of DRM regions from cultured cells contributed to the redistribution of γ-secretase components and Aß production. Taken together, the present analyses suggest that the lowered cholesterol content in DRM regions may be a cause of sporadic/late-onset AD by enhancing overall Aß generation.

Decrease in p3-Alcß37 and p3-Alcß40, products of Alcadein ß generated by γ-secretase cleavages, in aged monkeys and patients with Alzheimer's disease.

INTRODUCTION: Neuronal p3-Alcß peptides are generated from the precursor protein Alcadein ß (Alcß) through cleavage by α- and γ-secretases of the amyloid ß (Aß) protein precursor (APP). To reveal whether p3-Alcß is involved in Alzheimer's disease (AD) contributes for the development of novel therapy and/or drug targets. METHODS: We developed new sandwich enzyme-linked immunosorbent assay (sELISA) systems to quantitate levels of p3-Alcß in the cerebrospinal fluid (CSF). RESULTS: In monkeys, CSF p3-Alcß decreases with age, and the aging is also accompanied by decreased brain expression of Alcß. In humans, CSF p3-Alcß levels decrease to a greater extent in those with AD than in age-matched controls. Subjects carrying presenilin gene mutations show a significantly lower CSF p3-Alcß level. A cell study with an inverse modulator of γ-secretase remarkably reduces the generation of p3-Alcß37 while increasing the production of Aß42. DISCUSSION: Aging decreases the generation of p3-Alcß, and further significant decrease of p3-Alcß caused by aberrant γ-secretase activity may accelerate pathogenesis in AD.

Abnormal menstruation after radical trachelectomy and its effects on fertility.

AIM: To evaluate and identify the risk factors for abnormal menstruation after radical trachelectomy. METHODS: This study included 58 patients who underwent radical trachelectomy at our hospital between April 2005 and January 2018. Patients were divided into groups of those with no change in postoperative menstruation (regular [R] group; n = 46) and those with abnormal menstruation such as amenorrhea or hypomenorrhea (irregular [I] group; n = 12). The perioperative characteristics and fertility of the groups were compared retrospectively. The data were statistically analyzed using Student's t-test, Fisher's exact test and Mann-Whitney U test for univariate analysis and logistic regression analysis for multivariate analysis, with the level of statistical significance set at P < 0.05. RESULTS: Based on Federation of Gynecology and Obstetrics staging, 54 patients had stage IB1, 2 had stage IB2 and 2 had stage IIA1 cervical cancer. Eight patients received neoadjuvant chemotherapy. Pretreatment tumor size, residual uterine cavity length and neoadjuvant and postoperative chemotherapy use were not significantly different between the groups. Abnormal menstruation was significantly more common in patients with postoperative pelvic infection (R group, 13.0%; I group, 58.3%) and cervical stenosis (R group, 15.2%; I group, 58.3%). CONCLUSION: To maintain healthy menstruation even after radical trachelectomy, it is important to prevent postoperative pelvic infection and cervical stenosis.

Systemic Lupus Erythematosus Associated with Ovarian Cancer.

Systemic lupus erythematosus (SLE) may be associated with various types of malignancy. However, SLE occurring with ovarian cancer seems rare, and reliable therapeutic approaches for such cases have yet to be identified. We herein report a case of SLE with ovarian cancer that was successfully treated with corticosteroid, plasmapheresis and chemotherapy. This case may provide new insights into treatment approaches for SLE with ovarian cancer.

X11 and X11-like proteins regulate the level of extrasynaptic glutamate receptors.

X11/Mint 1 and X11-like (X11L)/Mint 2 are neuronal adaptor protein to regulate trafficking and/or localization of various membrane proteins. By analyzing the localization of neuronal membrane proteins in X11-, X11L-, and X11/X11L doubly deficient mice with membrane fractionation procedures, we found that deficient of X11 and X11L decreased the level of glutamate receptors in non-PSD fraction. This finding suggests that X11 and X11L regulate the glutamate receptor micro-localization to the extrasynaptic region. In vitro coimmunoprecipitation studies of NMDA receptors lacking various cytoplasmic regions with X11 and X11L proteins harboring domain deletion suggest that extrasynaptic localization of NMDA receptor may be as a result of the multiple interactions of the receptor subunits with X11 and X11L regulated by protein phosphorylation, while that of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor subunits is not dependent on the binding with X11 and X11L proteins. Because the loss of X11 and X11L tends to impair the exocytosis, but not endocytosis, of glutamate receptors, NMDA receptors are likely to be supplied to the extrasynaptic plasma membrane with a way distinct from the mechanism regulating the localization of NMDA receptors into synaptic membrane region. Reduced localization of NMDA receptor into the extrasynaptic region increased slightly the phosphorylation level of cAMP responsible element binding protein in brain of X11/X11L doubly deficient mice compare to wild-type mice, suggesting a possible role of X11 and X11L in the regulation of signal transduction pathway through extrasynaptic glutamate receptors. OPEN SCIENCE BADGES: This article has received a badge for *Open Materials* because it provided all relevant information to reproduce the study in the manuscript. The complete Open Science Disclosure form for this article can be found at the end of the article. More information about the Open Practices badges can be found at https://cos.io/our-services/open-science-badges/.

The cytoplasmic region of the amyloid ß-protein precursor (APP) is necessary and sufficient for the enhanced fast velocity of APP transport by kinesin-1.

Amyloid ß-protein precursor (APP) is transported mainly by kinesin-1 and at a higher velocity than other kinesin-1 cargos, such as Alcadein α (Alcα); this is denoted by the enhanced fast velocity (EFV). Interaction of the APP cytoplasmic region with kinesin-1, which is essential for EFV transport, is mediated by JNK-interacting protein 1 (JIP1). To determine the roles of interactions between the APP luminal region and cargo components, we monitored transport of chimeric cargo receptors, Alcα (luminal)-APP (cytoplasmic) and APP (luminal)-Alcα (cytoplasmic). Alcα-APP is transported at the EFV, whereas APP-Alcα is transported at the same velocity as wild-type Alcα. Thus, the cytoplasmic region of APP is necessary and sufficient for the EFV of APP transport by kinesin-1.

Induction of Amyloid-ß42 Production by Fipronil and Other Pyrazole Insecticides.

Generation of amyloid-ß peptides (Aßs) by proteolytic cleavage of the amyloid-ß protein precursor (AßPP), especially increased production of Aß42/Aß43 over Aß40, and their aggregation as oligomers and plaques, represent a characteristic feature of Alzheimer's disease (AD). In familial AD (FAD), altered Aß production originates from specific mutations of AßPP or presenilins 1/2 (PS1/PS2), the catalytic subunits of γ-secretase. In sporadic AD, the origin of altered production of Aßs remains unknown. We hypothesize that the 'human chemical exposome' contains products able to favor the production of Aß42/Aß43 over Aß40 and shorter Aßs. To detect such products, we screened a library of 3500 + compounds in a cell-based assay for enhanced Aß42/Aß43 production. Nine pyrazole insecticides were found to induce a ß- and γ-secretase-dependent, 3-10-fold increase in the production of extracellular Aß42 in various cell lines and neurons differentiated from induced pluripotent stem cells derived from healthy and FAD patients. Immunoprecipitation/mass spectrometry analyses showed increased production of Aßs cleaved at positions 42/43, and reduced production of peptides cleaved at positions 38 and shorter. Strongly supporting a direct effect on γ-secretase activity, pyrazoles shifted the cleavage pattern of another γ-secretase substrate, alcadeinα, and shifted the cleavage of AßPP by highly purified γ-secretase toward Aß42/Aß43. Focusing on fipronil, we showed that some of its metabolites, in particular the persistent fipronil sulfone, also favor the production of Aß42/Aß43 in both cell-based and cell-free systems. Fipronil administered orally to mice and rats is known to be metabolized rapidly, mostly to fipronil sulfone, which stably accumulates in adipose tissue and brain. In conclusion, several widely used pyrazole insecticides enhance the production of toxic, aggregation prone Aß42/Aß43 peptides, suggesting the possible existence of environmental "Alzheimerogens" which may contribute to the initiation and propagation of the amyloidogenic process in sporadic AD.

Phosphorylation of multiple sites within an acidic region of Alcadein α is required for kinesin-1 association and Golgi exit of Alcadein α cargo.

Alcadein α (Alcα) is a major cargo of kinesin-1 that is subjected to anterograde transport in neuronal axons. Two tryptophan- and aspartic acid-containing (WD) motifs located in its cytoplasmic domain directly bind the tetratricopeptide repeat (TPR) motifs of the kinesin light chain (KLC), which activate kinesin-1 and recruit kinesin-1 to Alcα cargo. We found that phosphorylation of three serine residues in the acidic region located between the two WD motifs is required for interaction with KLC. Phosphorylation of these serine residues may alter the disordered structure of the acidic region to induce direct association with KLC. Replacement of these serines with Ala results in a mutant that is unable to bind kinesin-1, which impairs exit of Alcα cargo from the Golgi. Despite this deficiency, the compromised Alcα mutant was still transported, albeit improperly by vesicles following missorting of the Alcα mutant with amyloid ß-protein precursor (APP) cargo. This suggests that APP partially compensates for defective Alcα in anterograde transport by providing an alternative cargo receptor for kinesin-1.

Phosphorylation of KLC1 modifies interaction with JIP1 and abolishes the enhanced fast velocity of APP transport by kinesin-1.

In neurons, amyloid ß-protein precursor (APP) is transported by binding to kinesin-1, mediated by JNK-interacting protein 1b (JIP1b), which generates the enhanced fast velocity (EFV) and efficient high frequency (EHF) of APP anterograde transport. Previously, we showed that EFV requires conventional interaction between the JIP1b C-terminal region and the kinesin light chain 1 (KLC1) tetratricopeptide repeat, whereas EHF requires a novel interaction between the central region of JIP1b and the coiled-coil domain of KLC1. We found that phosphorylatable Thr466 of KLC1 regulates the conventional interaction with JIP1b. Substitution of Glu for Thr466 abolished this interaction and EFV, but did not impair the novel interaction responsible for EHF. Phosphorylation of KLC1 at Thr466 increased in aged brains, and JIP1 binding to kinesin-1 decreased, suggesting that APP transport is impaired by aging. We conclude that phosphorylation of KLC1 at Thr466 regulates the velocity of transport of APP by kinesin-1 by modulating its interaction with JIP1b.

A case of laparoscopy-assisted vaginal cuff suturing for vaginal cuff dehiscence after total laparoscopic hysterectomy.

INTRODUCTION: Vaginal cuff dehiscence after hysterectomy is a rare complication and occurs in less than 1% of patients. It can present with serious complications, such as bowel evisceration and peritonitis. PRESENTATION OF CASE: A 51-year-old multigravida Korean woman underwent total laparoscopic hysterectomy for leiomyoma. Six months later, she reported lower abdominal pain and vaginal bleeding. Physical examination revealed rebound tenderness in the lower abdomen, and pelvic examination showed a small amount of vaginal bleeding with an evisceration of the small intestine through the vagina that exhibited healthy peristalsis. The eviscerated bowel, which seemed to be a part of the ileum, was carefully manually reduced transvaginally into the abdominal cavity. Laparoscopic observation revealed adhesions between the omentum, small intestine, and the peritoneum. Specifically, the small intestine was adhered around the vaginal cuff. An abdominal abscess was found in the left lower abdominal cavity. An adhesiotomy was performed and the abdominal abscess was removed and irrigated. Complete separation of the anterior and posterior vaginal cuff edges was obtained. The vaginal cuff was closed with interrupted 0-polydioxanone absorbable sutures without bowel injury. A 6-month follow-up examination revealed complete healing of the vaginal cuff. DISCUSSION: In this case, we were able to make use of both laparoscopic and transvaginal methods to perform a successful repair with a minimally invasive and safe technique. CONCLUSION: Laparoscopically assisted vaginal cuff suturing for vaginal cuff dehiscence after total laparoscopic hysterectomy was found to be effective, safe, and minimally invasive.

Facilitation of brain mitochondrial activity by 5-aminolevulinic acid in a mouse model of Alzheimer's disease.

The activities of mitochondrial enzymes, which are essential for neural function, decline with age and in age-related disease. In particular, the activity of cytochrome c oxidase (COX/complex IV) decreases in patients with Alzheimer's disease (AD). COX, a mitochondrial inner membrane protein complex that contains heme, plays an essential role in the electron transport chain that generates ATP. Heme synthesis begins with 5-aminolevulinic acid (5-ALA) in mitochondria. 5-ALA synthetase is the rate-limiting enzyme in heme synthesis, suggesting that supplementation with 5-ALA might help preserve mitochondrial activity in the aged brain. We administered a diet containing 5-ALA to triple-transgenic AD (3xTg-AD) model mice for 6 months, starting at 3 months of age. COX activity and protein expression, as well as mitochondrial membrane potential, were significantly higher in brains of 5-ALA-fed mice than in controls. Synaptotagmin protein levels were also significantly higher in 5-ALA-fed mice, suggesting improved preservation of synapses. Although brain Aß levels tended to decrease in 5-ALA-fed mice, we observed no other significant changes in other biochemical and pathological hallmarks of AD. Nevertheless, our study suggests that daily oral administration of 5-ALA could preserve mitochondrial enzyme activities in the brains of aged individuals, thereby contributing to the preservation of neural activity.

Alternative Selection of ß-Site APP-Cleaving Enzyme 1 (BACE1) Cleavage Sites in Amyloid ß-Protein Precursor (APP) Harboring Protective and Pathogenic Mutations within the Aß Sequence.

ß-Site APP-cleaving enzyme 1 (BACE1) cleaves amyloid ß-protein precursor (APP) at the bond between Met671 and Asp672 (ß-site) to generate the carboxyl-terminal fragment (CTFß/C99). BACE1 also cleaves APP at another bond between Thr681 and Gln682 (ß'-site), yielding CTFß'/C89. Cleavage of CTFß/C99 by γ-secretase generates Aß(1-XX), whereas cleavage of CTFß'/C89 generates Aß(11-XX). Thus, ß'-site cleavage by BACE1 is amyloidolytic rather than amyloidogenic. ß' cleavage of mouse APP is more common than the corresponding cleavage of human APP. We found that the H684R substitution within human Aß, which replaces the histidine in the human protein with the arginine found at the corresponding position in mouse, facilitated ß' cleavage irrespective of the species origin of BACE1, thereby significantly increasing the level of Aß(11-XX) and decreasing the level of Aß(1-XX). Thus, amino acid substitutions within the Aß sequence influenced the selectivity of alternative ß- or ß'-site cleavage of APP by BACE1. In familial Alzheimer's disease (FAD), the APP gene harbors pathogenic variations such as the Swedish (K670N/M671L), Leuven (E682K), and A673V mutations, all of which decrease Aß(11-40) generation, whereas the protective Icelandic mutation (A673T) increases generation of Aß(11-40). Thus, A673T promotes ß' cleavage of APP and protects subjects against AD. In addition, CTFß/C99 was cleaved by excess BACE1 activity to generate CTFß'/C89, followed by Aß(11-40), even if APP harbored pathogenic mutations. The resultant Aß(11-40) was more metabolically labile in vivo than Aß(1-40). Our analysis suggests that some FAD mutations in APP are amyloidogenic and/or amyloidolytic via selection of alternative BACE1 cleavage sites.