The role of the master cancer regulator Pin1 in the development and treatment of cancer.

This review examines the complex role of Pin1 in the development and treatment of cancer. Pin1 is the only peptidyl-prolyl isomerase (PPIase) that can recognize and isomerize phosphorylated Ser/Thr-Pro peptide bonds. Pin1 catalyzes a structural change in phosphorylated Ser/Thr-Pro motifs that can modulate protein function and thereby impact cell cycle regulation and tumorigenesis. The molecular mechanisms by which Pin1 contributes to oncogenesis are reviewed, including Pin1 overexpression and its correlation with poor cancer prognosis, and the contribution of Pin1 to aggressive tumor phenotypes involved in therapeutic resistance is discussed, with an emphasis on cancer stem cells, the epithelial-to-mesenchymal transition (EMT), and immunosuppression. The therapeutic potential of Pin1 inhibition in cancer is discussed, along with the promise and the difficulties in identifying potent, drug-like, small-molecule Pin1 inhibitors. The available evidence supports the efficacy of targeting Pin1 as a novel cancer therapeutic by analyzing the role of Pin1 in a complex network of cancer-driving pathways and illustrating the potential of synergistic drug combinations with Pin1 inhibitors for treating aggressive and drug-resistant tumors.

Interstitial macrophages are a focus of viral takeover and inflammation in COVID-19 initiation in human lung.

Early stages of deadly respiratory diseases including COVID-19 are challenging to elucidate in humans. Here, we define cellular tropism and transcriptomic effects of SARS-CoV-2 virus by productively infecting healthy human lung tissue and using scRNA-seq to reconstruct the transcriptional program in "infection pseudotime" for individual lung cell types. SARS-CoV-2 predominantly infected activated interstitial macrophages (IMs), which can accumulate thousands of viral RNA molecules, taking over 60% of the cell transcriptome and forming dense viral RNA bodies while inducing host profibrotic (TGFB1, SPP1) and inflammatory (early interferon response, CCL2/7/8/13, CXCL10, and IL6/10) programs and destroying host cell architecture. Infected alveolar macrophages (AMs) showed none of these extreme responses. Spike-dependent viral entry into AMs used ACE2 and Sialoadhesin/CD169, whereas IM entry used DC-SIGN/CD209. These results identify activated IMs as a prominent site of viral takeover, the focus of inflammation and fibrosis, and suggest targeting CD209 to prevent early pathology in COVID-19 pneumonia. This approach can be generalized to any human lung infection and to evaluate therapeutics.

Synergistic effects of using sodium hypochlorite (bleach) and desiccation in surface inactivation for Toxocara spp.

Toxocara cati and T. canis are parasitic nematodes found in the intestines of cats and dogs respectively, with a cosmopolitan distribution, and the potential for anthropozoonotic transmission, resulting in human toxocariasis. Spread of Toxocara spp. is primarily through the ingestion of embryonated eggs contaminating surfaces or uncooked food, or through the ingestion of a paratenic host containing a third-stage larva. The Toxocara spp. eggshell is composed of a lipid layer providing a permeability barrier, a chitinous layer providing structural strength, and thin vitelline and uterine layers, which combined create a biologically resistant structure, making the Toxocara spp. egg very hardy, and capable of surviving for years in the natural environment. The use of sodium hypochlorite, household bleach, as a disinfectant for Toxocara spp. eggs has been reported, with results varying from ineffective to limited effectiveness depending on parameters including contact time, concentration, and temperature. Desiccation or humidity levels have also been reported to have an impact on larval development and/or survival of Toxocara spp. eggs. However, to date, after a thorough search of the literature, no relevant publications have been found that evaluated the use of sodium hypochlorite and desiccation in combination. These experiments aim to assess the effects of using a combination of desiccation and 10% bleach solution (0.6% sodium hypochlorite) on fertilized or embryonated eggs of T. cati, T. canis, and T. vitulorum. Results of these experiments highlight the synergistic effects of desiccation and bleach, and demonstrate a relatively simple method for surface inactivation, resulting in a decrease in viability or destruction of T. cati, T. canis and T. vitulorum eggs. Implications for these findings may apply to larger scale elimination of ascarid eggs from both research, veterinary, and farming facilities to mitigate transmission.

Comparison of extracellular vesicle isolation methods for the study of exosome cargo within Toxocara canis and Toxocara cati excretory secretory (TES) products.

Toxocara is a genus of nematodes, which infects a variety of hosts, principally dogs and cats, with potential zoonotic risks to humans. Toxocara spp. larvae are capable of migrating throughout the host tissues, eliciting eosinophilic and granulomatous reactions, while surviving for extended periods of time, unchanged, in the host. It is postulated that larvae are capable of altering the host's immune response through the release of excretory-secretory products, containing both proteins and extracellular vesicles (EVs). The study of EVs has increased exponentially in recent years, largely due to their potential use as a diagnostic tool, and in molecular therapy. To this end, there have been multiple isolation methods described for the study of EVs. Here, we use nanoparticle tracking to compare the yield, size distribution, and % labelling of EV samples acquired through various reported methods, from larval cultures of Toxocara canis and T. cati containing Toxocara excretory-secretory products (TES). The methods tested include ultracentrifugation, polymer precipitation, magnetic immunoprecipitation, size exclusion chromatography, and ultrafiltration. Based on these findings, ultrafiltration produces the best results in terms of yield, expected particle size, and % labelling of sample. Transmission electron microscopy confirmed the presence of EVs with characteristic cup-shaped morphology. These findings can serve as a guide for those investigating EVs, particularly those released from multicellular organisms, such as helminths, for which few comparative analyses have been performed.

Mechanisms driving fasting-induced protection from genotoxic injury in the small intestine.

Genotoxic agents such as doxorubicin (DXR) can cause damage to the intestines that can be ameliorated by fasting. How fasting is protective and the optimal timing of fasting and refeeding remain unclear. Here, our analysis of fasting/refeeding-induced global intestinal transcriptional changes revealed metabolic shifts and implicated the cellular energetic hub mechanistic target of rapamycin complex 1 (mTORC1) in protecting from DXR-induced DNA damage. Our analysis of specific transcripts and proteins in intestinal tissue and tissue extracts showed that fasting followed by refeeding at the time of DXR administration reduced damage and caused a spike in mTORC1 activity. However, continued fasting after DXR prevented the mTORC1 spike and damage reduction. Surprisingly, the mTORC1 inhibitor, rapamycin, did not block fasting/refeeding-induced reduction in DNA damage, suggesting that increased mTORC1 is dispensable for protection against the initial DNA damage response. In Ddit4-/- mice [DDIT4 (DNA-damage-inducible transcript 4) functions to regulate mTORC1 activity], fasting reduced DNA damage and increased intestinal crypt viability vs. ad libitum-fed Ddit4-/- mice. Fasted/refed Ddit4-/- mice maintained body weight, with increased crypt proliferation by 5 days post-DXR, whereas ad libitum-fed Ddit4-/- mice continued to lose weight and displayed limited crypt proliferation. Genes encoding epithelial stem cell and DNA repair proteins were elevated in DXR-injured, fasted vs. ad libitum Ddit4-/- intestines. Thus, fasting strongly reduced intestinal damage when normal dynamic regulation of mTORC1 was lost. Overall, the results confirm that fasting protects the intestines against DXR and suggests that fasting works by pleiotropic - including both mTORC1-dependent and independent - mechanisms across the temporally dynamic injury response.NEW & NOTEWORTHY New findings are 1) DNA damage reduction following a 24-h fast depends on the timing of postfast refeeding in relation to chemotherapy initiation; 2) fasting/refeeding-induced upregulation of mTORC1 activity is not required for early (6 h) protection against DXR-induced DNA damage; and 3) fasting increases expression of intestinal stem cell and DNA damage repair genes, even when mTORC1 is dysregulated, highlighting fasting's crucial role in regulating mTORC1-dependent and independent mechanisms in the dynamic recovery process.

Cestodiasis in 2 Puerto Rican crested anoles.

Two adult male Puerto Rican crested anoles (Anolis cristatellus cristatellus) housed in a research facility were presented with debilitation and were euthanized. On autopsy, anole 1 had a large cystic white structure in the left pelvic limb, which protruded through the ruptured epidermis, and a large, poorly demarcated swelling in the right caudal abdomen. Anole 2 had masses in the mid-dorsum, caudal dorsum, left pelvic limb, and tail. These masses contained variably sized cestode larvae, which ruptured into the coelomic cavity. Evaluation of the larvae revealed a thickened and wrinkled anterior end, with a cleft-like invagination, consistent with either a plerocercoid sparganum or a tetrathyridium. Histologically, several cestode larvae were contained in the body wall of both anoles. These were up to 650 µm in diameter, with a thin tegument and a spongy parenchyma. The spongy parenchyma contained numerous, up to 30 µm diameter, sharply demarcated, basophilic-to-black structures (calcareous corpuscles). There was pneumonia and hepatitis in anole 2, suggestive of potential secondary infection subsequent to immunosuppression. Molecular amplification of the cytochrome C oxidase subunit 1 revealed 100% homology for the COX1 gene of the diphyllobothriid tapeworm Spirometra erinaceieuropaei, also known as Spirometra mansoni.

Evaluation of parasitic infections with histological and molecular detection in Anoles from St. Kitts.

Anoles are a widely distributed group of reptiles that are also increasing in popularity as a pet animal. These species have been described as paratenic and intermediate hosts for many parasites of veterinary and medical importance. However, while human and animals such as cats and dogs may often interact with anoles, little research has been conducted in terms of assessing the full potential of anoles to harbor parasites of public and veterinary health significance. The current study analyzed the parasites of anoles captured and dissected in St. Kitts, West Indies, to describe the parasitic population within these reptiles and evaluate the potential role of anoles as intermediate/paratenic hosts. From March until May of 2019, samples from 11 anoles were collected following humane euthanasia and evaluated grossly, histologically, and molecularly for the presence of parasites. Histologic evaluation revealed adult ascarids, oxyurids, and spirurids within the stomach, as well as intraluminal adult ascarids within the intestine, and encysted larval trematodes within the intestinal wall. Molecular analysis based on 18S rRNA and ITS2 DNA markers for nematodes and trematodes, respectively, identified Physaloptera sp. (99.4-99.8% identical with Physaloptera turgida, Physaloptera mirandai, Physaloptera retusa, Physaloptera rara) and Platynosomum illiciens (99.7% identical). Phylogenetic analysis of ITS 2 sequences of P. illiciens from this study cladded monophyletically with the same species from different geographic areas and hosts. Our study highlights the importance of understanding the role anoles play in the life cycles of less well-studied parasites, and the potential of these animals to act as intermediate or paratenic hosts.

Diagnosis of canine intestinal parasites: Improved detection of Dipylidium caninum infection through coproantigen testing.

Intestinal parasites, including cestodes like Dipylidium caninum, are common in dogs in the United States of America (USA), but fecal flotation consistently, and, at times, dramatically, fails to identify many of these infections. To determine the extent to which including coproantigen testing for D. caninum would improve the identification of dogs infected with this cestode, we evaluated fecal samples from 877 dogs (589 pet and 288 from municipal shelters) from six USA states using zinc sulfate (specific gravity 1.24) fecal flotation with centrifugation along with coproantigen detection for Giardia sp., hookworms, ascarids, and Trichuris vulpis. For D. caninum, PCR of perianal swabs was included. Intestinal parasite infections were identified, using centrifugal fecal flotation or coproantigen, in 265 dogs (13.2 % pet, 64.9 % shelter). Dipylidium caninum infection was detected in 5.6 % of dogs with the combination of coproantigen and centrifugal fecal flotation, and 7.3 % of dogs when perianal swab results were included; prevalence varied by diagnostic method, population, and geographic region. In pet dogs, D. caninum infection was identified by fecal flotation (0), coproantigen (2.2 %), or perianal swabs (1.2 %). The same methods revealed infection in 0.3 %, 12.5 %, and 11.1 % of shelter dogs, respectively. Frequent use of praziquantel in shelter dogs (116/288; 40.3 %) may have reduced prevalence. Positive and negative agreement of D. caninum coproantigen with perianal swab PCR in pet dogs was 85.7 % and 98.8 %, respectively. Multiple logistic regression analysis accounting for region, population, and age found D. caninum infection to be more common in shelter dogs relative to pet (adjusted OR 4.91 [2.48, 10.24]) and in the Southcentral and Southeast regions relative to North (adjusted OR 9.59 [1.92, 174.13] and 17.69 [3.67, 318.09] respectively). Coproantigen testing also enhanced the detection of other intestinal parasites over fecal flotation alone, including Giardia sp. (14.7 % vs 3.3 %), hookworms (13.8 % vs 8.4 %), ascarids (2.9 % vs 2.2 %), and T. vulpis (2.9 % vs 1.4 %). Together, these data indicate that the coproantigen assay employed increases detection of D. caninum infections several fold, supporting the use of this test in clinical practice, and add to a growing body of research documenting enhanced diagnosis through implementation of multiple laboratory-based methods.

DoSurvive: A webtool for investigating the prognostic power of a single or combined cancer biomarker.

We present DoSurvive, a user-friendly survival analysis web tool and a cancer prognostic biomarker centered database. DoSurvive is the first database that allows users to perform multivariant survival analysis for cancers with customized gene/patient list. DoSurvive offers three survival analysis methods, Log rank test, Cox regression and accelerated failure time model (AFT), for users to analyze five types of quantitative features (mRNA, miRNA, lncRNA, protein and methylation of CpG islands) with four survival types, i.e. overall survival, disease-specific survival, disease-free interval, and progression-free interval, in 33 cancer types. Notably, the implemented AFT model provides an alternative method for genes/features which failed the proportional hazard assumption in Cox regression. With the unprecedented number of survival models implemented and high flexibility in analysis, DoSurvive is a unique platform for the identification of clinically relevant targets for cancer researcher and practitioners. DoSurvive is freely available at http://dosurvive.lab.nycu.edu.tw/.

Abnormal Lymphatic Sphingosine-1-Phosphate Signaling Aggravates Lymphatic Dysfunction and Tissue Inflammation.

BACKGROUND: Lymphedema is a global health problem with no effective drug treatment. Enhanced T-cell immunity and abnormal lymphatic endothelial cell (LEC) signaling are promising therapeutic targets for this condition. Sphingosine-1-phosphate (S1P) mediates a key signaling pathway required for normal LEC function, and altered S1P signaling in LECs could lead to lymphatic disease and pathogenic T-cell activation. Characterizing this biology is relevant for developing much needed therapies. METHODS: Human and mouse lymphedema was studied. Lymphedema was induced in mice by surgically ligating the tail lymphatics. Lymphedematous dermal tissue was assessed for S1P signaling. To verify the role of altered S1P signaling effects in lymphatic cells, LEC-specific S1pr1-deficient (S1pr1LECKO) mice were generated. Disease progression was quantified by tail-volumetric and -histopathologic measurements over time. LECs from mice and humans, with S1P signaling inhibition, were then cocultured with CD4 T cells, followed by an analysis of CD4 T-cell activation and pathway signaling. Last, animals were treated with a monoclonal antibody specific to P-selectin to assess its efficacy in reducing lymphedema and T-cell activation. RESULTS: Human and experimental lymphedema tissues exhibited decreased LEC S1P signaling through S1P receptor 1 (S1PR1). LEC S1pr1 loss-of-function exacerbated lymphatic vascular insufficiency, tail swelling, and increased CD4 T-cell infiltration in mouse lymphedema. LECs, isolated from S1pr1LECKO mice and cocultured with CD4 T cells, resulted in augmented lymphocyte differentiation. Inhibiting S1PR1 signaling in human dermal LECs promoted T-helper type 1 and 2 (Th1 and Th2) cell differentiation through direct cell contact with lymphocytes. Human dermal LECs with dampened S1P signaling exhibited enhanced P-selectin, an important cell adhesion molecule expressed on activated vascular cells. In vitro, P-selectin blockade reduced the activation and differentiation of Th cells cocultured with shS1PR1-treated human dermal LECs. P-selectin-directed antibody treatment improved tail swelling and reduced Th1/Th2 immune responses in mouse lymphedema. CONCLUSIONS: This study suggests that reduction of the LEC S1P signaling aggravates lymphedema by enhancing LEC adhesion and amplifying pathogenic CD4 T-cell responses. P-selectin inhibitors are suggested as a possible treatment for this pervasive condition.

Abnormal lymphatic S1P signaling aggravates lymphatic dysfunction and tissue inflammation.

BACKGROUND: Lymphedema is a global health problem with no effective drug treatment. Enhanced T cell immunity and abnormal lymphatic endothelial cell (LEC) signaling are promising therapeutic targets for this condition. Sphingosine-1-phosphate (S1P) mediates a key signaling pathway required for normal LEC function, and altered S1P signaling in LECs could lead to lymphatic disease and pathogenic T cell activation. Characterizing this biology is relevant for developing much-needed therapies. METHODS: Human and mouse lymphedema was studied. Lymphedema was induced in mice by surgically ligating the tail lymphatics. Lymphedematous dermal tissue was assessed for S1P signaling. To verify the role of altered S1P signaling effects in lymphatic cells, LEC-specific S1pr1 -deficient ( S1pr1 LECKO ) mice were generated. Disease progression was quantified by tail-volumetric and -histopathological measurements over time. LECs from mice and humans, with S1P signaling inhibition, were then co-cultured with CD4 T cells, followed by an analysis of CD4 T cell activation and pathway signaling. Finally, animals were treated with a monoclonal antibody specific to P-selectin to assess its efficacy in reducing lymphedema and T cell activation. RESULTS: Human and experimental lymphedema tissues exhibited decreased LEC S1P signaling through S1PR1. LEC S1pr1 loss-of-function exacerbated lymphatic vascular insufficiency, tail swelling, and increased CD4 T cell infiltration in mouse lymphedema. LECs, isolated from S1pr1 LECKO mice and co-cultured with CD4 T cells, resulted in augmented lymphocyte differentiation. Inhibiting S1PR1 signaling in human dermal LECs (HDLECs) promoted T helper type 1 and 2 (Th1 and Th2) cell differentiation through direct cell contact with lymphocytes. HDLECs with dampened S1P signaling exhibited enhanced P-selectin, an important cell adhesion molecule expressed on activated vascular cells. In vitro , P-selectin blockade reduced the activation and differentiation of Th cells co-cultured with sh S1PR1 -treated HDLECs. P-selectin-directed antibody treatment improved tail swelling and reduced Th1/Th2 immune responses in mouse lymphedema. CONCLUSION: This study suggests that reduction of the LEC S1P signaling aggravates lymphedema by enhancing LEC adhesion and amplifying pathogenic CD4 T cell responses. P-selectin inhibitors are suggested as a possible treatment for this pervasive condition. Clinical Perspective: What is New?: Lymphatic-specific S1pr1 deletion exacerbates lymphatic vessel malfunction and Th1/Th2 immune responses during lymphedema pathogenesis. S1pr1 -deficient LECs directly induce Th1/Th2 cell differentiation and decrease anti-inflammatory Treg populations. Peripheral dermal LECs affect CD4 T cell immune responses through direct cell contact.LEC P-selectin, regulated by S1PR1 signaling, affects CD4 T cell activation and differentiation.P-selectin blockade improves lymphedema tail swelling and decreases Th1/Th2 population in the diseased skin.What Are the Clinical Implications?: S1P/S1PR1 signaling in LECs regulates inflammation in lymphedema tissue.S1PR1 expression levels on LECs may be a useful biomarker for assessing predisposition to lymphatic disease, such as at-risk women undergoing mastectomyP-selectin Inhibitors may be effective for certain forms of lymphedema.

Functional screening of lysosomal storage disorder genes identifies modifiers of alpha-synuclein neurotoxicity.

Heterozygous variants in the glucocerebrosidase (GBA) gene are common and potent risk factors for Parkinson's disease (PD). GBA also causes the autosomal recessive lysosomal storage disorder (LSD), Gaucher disease, and emerging evidence from human genetics implicates many other LSD genes in PD susceptibility. We have systemically tested 86 conserved fly homologs of 37 human LSD genes for requirements in the aging adult Drosophila brain and for potential genetic interactions with neurodegeneration caused by α-synuclein (αSyn), which forms Lewy body pathology in PD. Our screen identifies 15 genetic enhancers of αSyn-induced progressive locomotor dysfunction, including knockdown of fly homologs of GBA and other LSD genes with independent support as PD susceptibility factors from human genetics (SCARB2, SMPD1, CTSD, GNPTAB, SLC17A5). For several genes, results from multiple alleles suggest dose-sensitivity and context-dependent pleiotropy in the presence or absence of αSyn. Homologs of two genes causing cholesterol storage disorders, Npc1a / NPC1 and Lip4 / LIPA, were independently confirmed as loss-of-function enhancers of αSyn-induced retinal degeneration. The enzymes encoded by several modifier genes are upregulated in αSyn transgenic flies, based on unbiased proteomics, revealing a possible, albeit ineffective, compensatory response. Overall, our results reinforce the important role of lysosomal genes in brain health and PD pathogenesis, and implicate several metabolic pathways, including cholesterol homeostasis, in αSyn-mediated neurotoxicity.

Tau polarizes an aging transcriptional signature to excitatory neurons and glia.

Aging is a major risk factor for Alzheimer's disease (AD), and cell-type vulnerability underlies its characteristic clinical manifestations. We have performed longitudinal, single-cell RNA-sequencing in Drosophila with pan-neuronal expression of human tau, which forms AD neurofibrillary tangle pathology. Whereas tau- and aging-induced gene expression strongly overlap (93%), they differ in the affected cell types. In contrast to the broad impact of aging, tau-triggered changes are strongly polarized to excitatory neurons and glia. Further, tau can either activate or suppress innate immune gene expression signatures in a cell-type-specific manner. Integration of cellular abundance and gene expression pinpoints nuclear factor kappa B signaling in neurons as a marker for cellular vulnerability. We also highlight the conservation of cell-type-specific transcriptional patterns between Drosophila and human postmortem brain tissue. Overall, our results create a resource for dissection of dynamic, age-dependent gene expression changes at cellular resolution in a genetically tractable model of tauopathy.

Taenia crassiceps Cysticercosis in a Wild Muskrat and a Domestic Dog in the Northeastern United States.

Taenia crassiceps is a parasite of wild canids and dogs that serve as definite hosts, harboring the adult cestode, whereas rodents are the intermediate hosts in which the metacestode/cysticercus/larval stage occurs. Fecal-oral transmission ensures the parasite's lifecycle. At times, dogs and humans act as accidental intermediate hosts. Despite the public health concern this parasite warrants, its epidemiology remains unclear. In this report, we document the occurrence of metacestodes of T. crassiceps in a muskrat (Ondatra zibethicus) and a domestic dog from the northeastern United States, a development that necessitates increased awareness and surveillance to tackle this disease of "one health" significance. Taenia crassiceps cysticercosis was confirmed in an adult male muskrat in February 2018 and in a 4-year-old female spayed Staffordshire Bull Terrier in December 2020. Parasitological and histopathologic examination of both cases revealed cysticerci with the characteristic rostellar hook morphology that aided in Taenia species identification. In the muskrat case specifically, partial sequencing of the mitochondrial cytochrome oxidase gene confirmed the species identity as T. crassiceps. We report T. crassiceps occurrence in a muskrat in New York State for the first time and document a case presentation in a domestic dog from New Jersey that was infected with metacestode stages of this parasite. Given the detection of this parasite in the northeastern United States, T. crassiceps infection, which otherwise is considered a rare disease, should be on the radar of veterinary, medical and wildlife biologists for timely diagnosis and interventions.

Neuroendocrinology of the lung revealed by single-cell RNA sequencing.

Pulmonary neuroendocrine cells (PNECs) are sensory epithelial cells that transmit airway status to the brain via sensory neurons and locally via calcitonin gene-related peptide (CGRP) and γ- aminobutyric acid (GABA). Several other neuropeptides and neurotransmitters have been detected in various species, but the number, targets, functions, and conservation of PNEC signals are largely unknown. We used scRNAseq to profile hundreds of the rare mouse and human PNECs. This revealed over 40 PNEC neuropeptide and peptide hormone genes, most cells expressing unique combinations of 5-18 genes. Peptides are packaged in separate vesicles, their release presumably regulated by the distinct, multimodal combinations of sensors we show are expressed by each PNEC. Expression of the peptide receptors predicts an array of local cell targets, and we show the new PNEC signal angiotensin directly activates one subtype of innervating sensory neuron. Many signals lack lung targets so may have endocrine activity like those of PNEC-derived carcinoid tumors. PNECs are an extraordinarily rich and diverse signaling hub rivaling the enteroendocrine system.