Rise in calcium (Ca2+) and hyperactivation of Ca2+-dependent phosphatase calcineurin are two key determinants of α-synuclein (α-syn) pathobiology implicated in synucleinopathies such as Dementia with Lewy Bodies and Parkinson's Disease. Calcineurin activity can be inhibited with FK506, a Food and Drug Administration (FDA)-approved compound. Our previous work demonstrated a protective effect of sub-saturating doses of FK506 against α-syn pathology in a rat model of α-syn neurodegeneration. This neuroprotection was associated with the phosphorylation of GAP-43. In this study, we present evidence that phosphorylation of GAP-43 regulated by calcineurin is a critical determinant for neurite branching and synapse formation. Phosphorylation of GAP-43 promotes neurite branching and synapse formation whereas dephosphorylation prevents it. Therefore, our findings provide a novel way in which GAP-43 activity can be regulated by calcineurin and provide the mechanistic basis for neuroprotection for FK506 effects in neurons experiencing α-syn proteotoxic stress. ### Competing Interest Statement The authors have declared no competing interest.

Introduction:Elevated calcium (Ca2+) levels and hyperactivation of the Ca2+-dependent phosphatase calcineurin are key factors in α-synuclein (α-syn) pathobiology in Dementia with Lewy Bodies and Parkinson's Disease (PD). Calcineurin activity can be inhibited by FK506, an FDA-approved compound. Our previous work demonstrated that sub-saturating doses of FK506 provide neuroprotection against α-syn pathology in a rat model of α-syn neurodegeneration, an effect associated with the phosphorylation of growth-associated protein 43 (GAP-43). Methods:To investigate the role of GAP-43 phosphorylation, we generated phosphomutants at the calcineurin-sensitive sites and expressed them in PC12 cells and primary rat cortical neuronal cultures to assess their effects on neurite morphology and synapse formation. Additionally, we performed immunoprecipitation mass spectrometry in HeLa cells to identify binding partners of these phosphorylation sites. Finally, we evaluated the ability of these phosphomutants to modulate α-syn toxicity. Results:In this study, we demonstrate that calcineurin-regulated phosphorylation at S86 and T172 of GAP-43 is a crucial determinant of neurite branching and synapse formation. A phosphomimetic GAP-43 mutant at these sites enhances both processes and provides protection against α-syn-induced neurodegeneration. Conversely, the phosphoablative mutant prevents neurite branching and synapse formation while exhibiting increased interactions with ribosomal proteins. Discussion:Our findings reveal a novel mechanism by which GAP-43 activity is regulated through phosphorylation at calcineurin-sensitive sites. These findings suggest that FK506's neuroprotective effects may be partially mediated through GAP-43 phosphorylation, providing a potential target for therapeutic intervention in synucleinopathies.

Actin-mediated mitochondrial fission is essential for cellular homeostasis, yet the mechanisms by which actin is recruited to mitochondria and how it couples the outer and inner mitochondrial membranes (IMM) remain poorly understood. Using a phosphoproteomic screen in a rat model of α-synucleinopathy, we identified BASP1 as a calcineurin-dependent substrate that is constitutively dephosphorylated under pathological Ca2+ elevations and phosphorylated under neuroprotective calcineurin inhibition. Immunoprecipitation and mass spectrometry of phosphomutant BASP1 expressed in neurons revealed that dephosphorylation promotes interactions with actin and IMM proteins. Dephosphorylated BASP1 recruits actin to mitochondria, while subsequent phosphorylation enables actin-mediated mitochondrial fission and neurite elongation. Constitutive dephosphorylation, as it occurs in α-synucleinopathy, impairs mitochondrial fission, inhibits neurite growth and promotes α-synuclein aggregation. Our findings position BASP1 as a Ca2+-CaN-regulated hub that coordinates actin remodeling and couples mitochondrial membranes to drive fission, revealing a mechanistic axis linking mitochondrial dysfunction to neuronal morphogenesis and α-synuclein pathobiology. ### Competing Interest Statement The authors have declared no competing interest. Parkinson's Foundation, https://ror.org/05mx85j86, PF-JFA-1949

Introduction: Rise in Calcium (Ca2+) and hyperactive Ca2+-dependent phosphatase calcineurin represent two key determinants of a-synuclein (a-syn) pathobiology implicated in Parkinson’s Disease (PD) and other neurodegenerative diseases. Calcineurin activity can be inhibited with FK506, a Food and Drug Administration (FDA)-approved compound. Our previous work demonstrated a protective effect of low doses of FK506 against a-syn pathology in various models of a-syn related pathobiology.Methods: Control and a-syn-expressing mice (12-18 months old) were injected with vehicle or two single doses of FK506 administered 4 days apart. Cerebral cortex and serum from these mice were collected and assayed using a meso scale discovery quickplex SQ 120 for cytokines and Enzyme-linked immunosorbent assay for IGF-1.Results: In this study we present evidence that reducing calcineurin activity with FK506 in a-syn transgenic mice increased insulin growth factor (IGF-1), while simultaneously decreasing IL-2 levels in both cerebral cortex and serum.Discussion: The highly conserved Ca2+/calcineurin signaling pathway is known to be affected in a-syn-dependent human disease. FK506, an already approved drug for other uses, exhibits high brain penetrance and a proven safety profile. IL-2 and IGF-1 are produced throughout life and can be measured using standard clinical methods. Our findings provide two potential biomarkers that could guide a clinical trial of FK506 in PD patients, without posing significant logistical or regulatory challenges.

Octopamine is a well-established invertebrate neurotransmitter involved in fight or flight responses. In mammals, its function was replaced by epinephrine. Nevertheless, it is present at trace amounts and can modulate the release of monoamine neurotransmitters by a yet unidentified mechanism. Here, through a multidisciplinary approach utilizing in vitro and in vivo models of α-synucleinopathy, we uncovered an unprecedented role for octopamine in driving the conversion from toxic to neuroprotective astrocytes in the cerebral cortex by fostering aerobic glycolysis. Physiological levels of neuron-derived octopamine act on astrocytes via a trace amine-associated receptor 1–Orai1–Ca 2+ –calcineurin-mediated signaling pathway to stimulate lactate secretion. Lactate uptake in neurons via the monocarboxylase transporter 2–calcineurin-dependent pathway increases ATP and prevents neurodegeneration. Pathological increases of octopamine caused by α-synuclein halt lactate production in astrocytes and short-circuits the metabolic communication to neurons. Our work provides a unique function of octopamine as a modulator of astrocyte metabolism and subsequent neuroprotection with implications to α-synucleinopathies.

Epidemiologic studies in diabetic patients as well as research in model organisms have indicated the potential of metformin as a drug candidate for the treatment of various types of cancer, including breast cancer. To date most of the anti-cancer properties of metformin have, in large part, been attributed either to the inhibition of mitochondrial NADH oxidase complex (Complex I in the electron transport chain) or the activation of AMP-activated kinase (AMPK). However, it is becoming increasingly clear that AMPK activation may be critical to alleviate metabolic and energetic stresses associated with tumor progression suggesting that it may, in fact, attenuate the toxicity of metformin instead of promoting it. Here, we demonstrate that AMPK opposes the detrimental effects of mitochondrial complex I inhibition by enhancing glycolysis at the expense of, and in a manner dependent on, pyruvate availability. We also found that metformin forces cells to rewire their metabolic grid in a manner that depends on AMPK, with AMPK-competent cells upregulating glycolysis and AMPK-deficient cell resorting to ketogenesis. In fact, while the killing effects of metformin were largely rescued by pyruvate in AMPKcompetent cells, AMPK-deficient cells required instead acetoacetate, a product of fatty acid catabolism indicating a switch from sugar to fatty acid metabolism as a central resource for ATP production in these cells. In summary, our results indicate that AMPK activation is not responsible for metformin anticancer activity and may instead alleviate energetic stress by activating glycolysis.

Ykt6 is a soluble N-ethylmaleimide sensitive factor activating protein receptor (SNARE) critically involved in diverse vesicular fusion pathways. While most SNAREs rely on transmembrane domains for their activity, Ykt6 dynamically cycles between the cytosol and membrane-bound compartments where it is active. The mechanism that regulates these transitions and allows Ykt6 to achieve specificity toward vesicular pathways is unknown. Using a Parkinson's disease (PD) model, we found that Ykt6 is phosphorylated at an evolutionarily conserved site which is regulated by Ca2+ signaling. Through a multidisciplinary approach, we show that phosphorylation triggers a conformational change that allows Ykt6 to switch from a closed cytosolic to an open membrane-bound form. In the phosphorylated open form, the spectrum of protein interactions changes, leading to defects in both the secretory and autophagy pathways, enhancing toxicity in PD models. Our studies reveal a mechanism by which Ykt6 conformation and activity are regulated with potential implications for PD.

Neurotropic alphaherpesviruses initiate infection in exposed mucosal tissues and, unlike most viruses, spread rapidly to sensory and autonomic nerves where life-long latency is established1. Recurrent infections arise sporadically from the peripheral nervous system throughout the life of the host, and invasion of the central nervous system may occur, with severe outcomes2. These viruses directly recruit cellular motors for transport along microtubules in nerve axons, but how the motors are manipulated to deliver the virus to neuronal nuclei is not understood. Here, using herpes simplex virus type I and pseudorabies virus as model alphaherpesviruses, we show that a cellular kinesin motor is captured by virions in epithelial cells, carried between cells, and subsequently used in neurons to traffic to nuclei. Viruses assembled in the absence of kinesin are not neuroinvasive. The findings explain a critical component of the alphaherpesvirus neuroinvasive mechanism and demonstrate that these viruses assimilate a cellular protein as an essential proviral structural component. This principle of viral assimilation may prove relevant to other virus families and offers new strategies to combat infection.

Introduction: In our previous studies it was concluded that the zinc (Zn) level in in human expressed prostatic fluid (EPF) is a first candidate with the role of offering a new, simple, fast, reliable, and non-invasive diagnostic tool for prostate cancer (PCa) screening. However it was unclear how other non-prostatic diseases inherent of the old persons, including heart diseases, atherosclerosis, diabetes, asymptomatic cancer of different localization and some others, impact on a chemical element composition of expressed prostatic fluid. Thus, the purpose of this study was to evaluate whether significant changes in the levels of Zn in EPF exist in patients with bladder cancer (BC). Methods: Prostatic fluid levels of Zn were prospectively evaluated in 17 patients with BC and 51 healthy male inhabitants (control group). Measurements were performed using 109Cd radionuclide-induced energy dispersive X-ray fluorescent microanalysis developed by us. Results: Mean value  standard deviation of mean (MSD) for concentration of Zn in the EPF of healthy males and patients with BC was 573202 mg/L and 625±108 mg/L, respectively. Using both parametric Student’s t-test and non-parametric Wilcoxon-Mann-Whitney U-test it was shown that the Zn concentration in the EPF of patients with BC did not differ from that in healthy subjects. Conclusion: Such serious illness as BC did not impact on the Zn concentration in the EPF.

Background : Prostate cancer (PCa) is an important and universal health problem of the man, particularly in developed countries. Prostate specific antigen (PSA) levels in blood do not provide the high reliability and precision required for an accurate screening for PCa. There is a need in a simple, rapid, direct, preferably non-invasive, and highly accurate biomarker and procedure for the screening for PCa. Objective : To report the results from a Russian population-based randomized study of a screening using the zinc (Zn) level in expressed prostatic fluid (EPF) to detect PCa. Methods : A total of 578 Russian ambulatory males aged over 50 years had their Zn levels in EPF determined. Men with their EPF Zn levels < 100 mg/L were subjected to further clinical examination including digital rectal examination (DRE) and transrectal prostate biopsy. Results : The Zn levels in EPF (mean ± standard deviation, SD) in all the men (n=578) was 618±377 mg/L and, for those of them with PCa (n=3), the mean Zn level was 15±20 mg/L ( p ≤0.001). The Zn level in EPF increased with age up to 70 years but over 71 years old this metal’s concentration began to decrease. In persons aged 51-60, 61-70, 71-80, and above 81 years, the mean ± SD values of the Zn levels in EPF were 602±329 (n=135), 655±395 (n=324), 539±354 (n=103), and 455±432 (n=16) mg/L, respectively. Among all the screened men, 33 (5.7%) had a Zn concentration in EPF < 100 mg/L. Of the patients who underwent prostate biopsy (n=11), 3 had PCa (positive predictive value, 27.3%). Conclusion : The usefulness of Zn levels in EPF as a reliable biomarker for the screening for PCa needs to be confirmed in other randomized trials.

Among different bio dosimeters, there is a need in methods that can rapidly and accurately determine individual exposure to ionizing radiation during examination a big cohort of people, particularly men.In present study contents of trace elements in the prostatic fluid were investigated to search for a new biological dosimeter.Prostatic fluid samples were obtained from 15 men aged 38 -77 years suffering from bladder cancer.All patients were treated with external (distant) gamma-ray therapy.Fractionated tumour irradiation was used with a total dose of 65 Gy.The tumour was daily irradiated with the dose ranged from 6 to 12 Gy.The prostate radiation dose was calculated according to the distribution of isodose levels within patient's body during the simulation of external local gamma-ray irradiation of bladder tumour.Transrectal digital massage was used to obtain expressed prostatic fluid samples before and 2 -4 times during radiation therapy for the energy dispersive X-ray fluorescent analysis.A day after the first irradiation of the prostate with the average dose 4.6 Gy to prostate concentration of zinc in the prostatic fluid was about three times lower the metal level before gamma-ray therapy.The dose detection threshold was calculated up to 0.1 Gy.The exponential function was the best for approximation the "dose-effect" dependence of the zinc level in prostatic fluid from the radiation exposure during external gamma-ray therapy of patients with bladder cancer.

The effect of age on Br, Fe, Rb, Sr, and Zn concentrations as well as on Zn/Br, Zn/Fe, Zn/Rb, and Zn/Sr content ratios in human prostatic fluid was investigated by 109Cd radionuclide-induced energy dispersive X-ray fluorescent microanalysis. Specimens of expressed prostatic fluid were obtained from 51 men (mean age 51 years, range 18-82 years) with apparently normal prostates using standard rectal massage procedure. Mean values (M ± SΕΜ) for concentration of trace elements (mg·L-1) in human prostate fluid were: Br 3.62±0.58, Fe 9.04±1.21, Rb 1.10±0.08, Sr 1.19±0.14, and Zn 573±28. Mean values for ratios of trace elements in human prostate fluid were: Zn/Br 523±103, Zn/Fe 105±16, Zn/Rb 661±63, and Zn/Sr 719±95. An age-related increase in Zn content and decrease in Br and Fe concentration was found. Moreover, the strongly pronounced increase in Zn/Br and Zn/Fe ratios was also observed.

Aim: Benign prostatic hyperplasia and prostate cancer is an internationally important health problem of the man, particularly in developed countries. The aim of this exploratory study was to evaluate whether significant difference in the levels of Zn and some other trace elements of prostatic fluid exist between the inflamed and malignantly transformed prostate. Methods: Prostatic fluid levels of Br, Fe, Rb, Sr, and Zn were prospectively evaluated in 52 patients with benign prostatic hyperplasia and 24 patients with prostate cancer. Measurements were performed using 109Cd radionuclide-induced energy dispersive X-ray fluorescent microanalysis. Prostatic fluid samples were divided into two portions. One was used for cytological study to exclude prostatitis, while the other was intended for trace element analysis. Results: Mean values ± standard error of means for concentration (mg/L) of trace element in the prostatic fluid of hyperplastic prostate were: Br 2.32 ± 0.30, Fe 11.5 ± 1.8, Rb 1.70 ± 0.23, Sr 1.41 ± 0.26, and Zn 488 ± 42. The contents of Rb and Zn were significantly lower (approximately 3.2 and 7.7 times, respectively) in fluid of cancerous prostate compared with those in fluid of hyperplastic prostate. Conclusion: There are significant differences in trace element contents in the fluid of hyperplastic and malignantly transformed prostate. The great decrease in levels of Rb and Zn in the fluid of cancerous prostate might demonstrate an involvement of these trace elements in etiology and pathogenesis of malignant prostate tumors. It was supposed that the differences in Rb and Zn levels in prostatic fluid can be used as tumor markers.

Background and objective: the prostate gland is subject to various disorders and of them chronic prostatitis (CP) is a complex disease. Various studies indicate some discrepancies regarding zinc (Zn) levels in prostatic fluid of normal and inflamed glands. The present study evaluated by systematic analysis the published data for Zn levels analyzed in expressed prostatic fluid (EPF) of normal and inflamed glands. Methods: The present systematic analysis included 25 studies, all of which were published in the years from 1961 to 2018 and selected by searching the databases Scopus, PubMed, MEDLINE, ELSEVIER-EMBASE, Cochrane Library, and the Web of Science. The articles were analyzed and “Median of Means” and “Range of Means” were used to examine heterogeneity of Zn concentrations in two groups of subjects – apparently healthy men “N” and patients with prostatitis “P”. Moreover, using the ratios of prostatic fluid Zn in “P” group to prostatic fluid Zn in “N” group (ZnP/ZnN) obtained (or calculated by us) in the reviewed studies, “Median of Means” and “Range of Means” for these ratios were found. The objective analysis was performed on data from the 25 studies, with total 285 subjects in “P” group and more than 900 subjects in “N” group. Results: The range of means of Zn concentration reported in the literature for normal EPF varies widely from 47.1 mg/L to 825 mg/L with median of means 501 mg/L. The range of means of Zn concentration for EPF of untreated prostatitis varies also widely from 56 mg/L to 491 mg/L, with median of means 268 mg/L. Thus, the obtained median of means for Zn concentration in normal human prostatic fluid is about two times higher than median of mean values of the element content in EPF of inflamed prostate. In other words, the analysis of 25 studies with discordant data regarding prostatic fluid concentration of Zn demonstrated that there is a significantly diminished concentration of Zn in EPF of patients with prostatitis compared to controls. Conclusion: There is a significant relationship between lowered Zn concentrations in prostatic fluid and prostatitis, but because of small sample size and high data heterogeneity, we recommend other primary studies.

Introduction: Benign prostatic hyperplasia (BPH) is a benign tumor that infects almost all men as they get older. To date, we still have no precise knowledge of the biochemical, cellular and molecular processes underlying the pathogenesis of BPH. One of the main functions of prostate gland is a production of prostatic fluid with extremely high concentration of Zn and elevated level some other TE and electrolytes. The aim of this exploratory study was to evaluate changes in the levels of Zn/Br, Zn/Fe, Zn/Rb, and Zn/Sr concentration ratio in prostatic fluid of the hyperplastic prostate. Methods: Prostatic fluid levels of Br, Fe, Rb, Sr, and Zn were prospectively evaluated in 52 patients with BPH and 38 healthy male inhabitants. Measurements were performed using 109Cd radionuclide-induced energy dispersive X-ray fluorescent microanalysis. Using obtained results the individual values of Zn/Br, Zn/Fe, Zn/Rb, and Zn/Sr concentration ratio were calculated. Results: Mean values ± standard error of means for Zn/Br, Zn/Fe, Zn/Rb, and Zn/Sr concentration ratio in the prostatic fluid of normal prostate were: 639 ± 122, 120 ± 19, 637 ± 69, and 733 ± 116, respectively. It was observed that in the secret of hyperplastic prostate ratios of Zn/Br, Zn/Fe, and Zn/Rb some decreased in a comparison with those in the secret of normal prostate however the found differences were not statistically significant. Conclusions: The alterations in levels of Zn and Rb in the EPF of hyperplastic prostate might demonstrate an involvement of these trace elements in etiology and pathogenesis of BPH. In spite of opposite directions in changes of Zn and Rb contents in EPF of BPH prostate the alteration in level of Zn/Rb ratio was not statistically significant. In our opinion, found changes in TE contents and their ratios in EPF of BPH prostate could indicate a suppression of specific functional activity of the gland.

The prevalence of thyroid dysfunction, including goiter and cancer, is higher in the elderly as compared to the younger population. An excess or deficiency of specific chemical element contents in thyroid parenchyma plays an important role ingoitroand carcinogenesis of this gland. The variation with age of the mass fraction of twenty elements (Al, B, Ba, Br, Ca, Cl, Cu, Fe, I, K, Li, Mg, Mn, Na, P, S, Si, Sr, V, and Zn) in normal thyroids of 33 females (mean age 54.5 years, range 3.5-87) was investigated. Measurements were performed using a combination of non-destructive and destructive methods: instrumental neutron activation analysis and inductively coupled plasma atomic emission spectrometry, respectively. Tissue samples were divided into two portions. One was used for morphological study while the other was intended for chemical element analysis. This work revealed that there is a statistically significant increase in Al, Br, Ca, Cu, Fe, P, S and Zn mass fraction, as well as a decrease in the Cl mass fraction in the normal thyroids of females during a lifespan. Results of the study showed that for elderly females there is a goitrogenic and carcinogenic association with elevated levels of the thyroid parenchymal chemical elements as Al, Br, Ca, Cu, Fe, and Zn, which increase intra-thyroidal oxidative stress.

The Journal of Cancer Metastasis and Treatment is an open access journal focused on cancer metastasis and treatment, including the occurrence, development, progression, metastasis, and treatment of oncologic disease. It covers basic, translational and clinical research related to cancer cell biology, genomics, precision medicine, oncology internal medicine, radiotherapy and radiology, obstetrics and gynecology, pediatrics, surgery, hematology, neurooncology, etc.

It is unclear why a prevalence of thyroid dysfunction is higher in the elderly as compared to the younger population. An excess or deficiency of trace element contents in thyroid may play important role in goitre- and carcinogenesis of gland.