Modulation of regenerative responses by retinoic and ascorbic acids in human apical papilla cells.

OBJECTIVE: This study investigated the bioactive effects of retinoic acid and ascorbic acid on hSCAPs in vitro. DESIGN: Cells were obtained from human third molars (n=4) and characterized for mesenchymal stem cell markers by flow cytometry. The experimental groups: control (α-MEM); vehicle control group (α-MEM + 0.17 % DMSO); retinoic acid 0.1, 1, and 10 µM; and ascorbic acid 3, 30, and 300 µM (n=8) were tested for cell viability (alamarBlue; 1, 3, and 7 days), total collagen synthesis (Sirius Red; 1 and 7 days), mineralized matrix formation (Alizarin red; 14 days), and the regulation of gene expression related to mineralization (ALPL and DSPP), cell migration (ITGAV and CXCL12) angiogenesis (VEGFA) and collagen synthesis (COL1A1 and COL3A1; RT-qPCR) on 1 and 7 days. ACTB and GAPDH were used as reference genes. Data were analyzed by ANOVA and complementary tests at a 5 % significance level. RESULTS: Ascorbic acid 300 µM increased viability, and retinoic acid reduced it dose-dependently. Retinoic acid 0.1 µM and ascorbic acid 30 and 300 µM increased mineralized matrix formation and total collagen synthesis, and retinoic acid 10 µM decreased. On day 1, 0.1 µM retinoic acid upregulated the gene expression of COL1A1, COL3A1, VEGFA, CXCL12, ALPL, DSPP e ITGAV, and 300 µM ascorbic acid upregulated COL1A1, COL3A1 and DSPP. However, on day 7, retinoic acid downregulated ALPL, COL3A1, CXCL12, and VEGFA and downregulated ITGAV and VEGFA. CONCLUSION: Retinoic acid 0.1 µM and ascorbic acid 300 µM biostimulated hSCAPs to differentiate into pro-regenerative phenotypes with potential application for REPs.

Single-cell RNA sequencing data locate ALDH1A2-mediated retinoic acid synthetic pathway to glomerular parietal epithelial cells.

Aldehyde dehydrogenase 1, family member A2, is a retinoic acid-synthesizing enzyme encoded by Aldh1a2 in mice and ALDH1A2 in humans. This enzyme is indispensable for kidney development, but its role in kidney physiology and pathophysiology remains to be fully defined. In this review, we mined single-cell and single-nucleus RNA sequencing databases of mouse and human kidneys and found that glomerular parietal epithelial cells (PECs) express a full set of genes encoding proteins needed for cellular vitamin A uptake, intracellular transport, and metabolism into retinoic acid. In particular, Aldh1a2/ALDH1A2 mRNAs are selectively enriched in mouse and human PECs. Aldh1a2 expression in PECs is greatly increased in a mouse model of anti-glomerular basement membrane glomerulonephritis and moderately induced in a mouse model of ischemia-reperfusion acute kidney injury. Aldh1a2 expression in PECs is substantially repressed in a chronic kidney disease mouse model combining diabetes, hypertension, and partial nephrectomy and is moderately repressed in mouse models of focal segmental glomerulosclerosis and diabetic nephropathy. Single-nucleus RNA sequencing data show that ALDH1A2 mRNA expression in PECs is diminished in patients with chronic kidney disease associated with diabetes, hypertension and polycystic kidney disease. In addition to data mining, we also performed Spearman's rank correlation coefficient analyses and identified gene transcripts correlated with Aldh1a2/ALDH1A2 transcripts in mouse PECs and PEC subtypes, and in human PECs of healthy subjects and patients with AKI or CKD. Furthermore, we conducted Gene Ontology pathway analyses and identified the biological pathways enriched among these Aldh1a2/ALDH1A2-correlated genes. Our data mining and analyses led us to hypothesize that ALDH1A2-mediated retinoic acid synthesis in PECs plays a yet-undefined role in the kidney and that its dysregulation mediates injury. Conditional, PEC-selective Aldh1a2 knockout, RNA silencing and transgenic mouse models will be useful tools to test this hypothesis. Clinical studies on genetics, epigenetics, expression and functions of ALDH1A2 and other genes needed for retinoic acid biosynthesis and signaling are also warranted.

Retinoic Acid Improves Vascular Endothelial Dysfunction by Inhibiting PI3K/AKT/YAP-Mediated Ferroptosis in Diabetes Mellitus.

BACKGROUND: Vascular endothelial dysfunction is the initial factor involved in cardiovascular injury in patients with diabetes. Retinoic acid is involved in improving vascular complications in patients with diabetes, but its protective mechanism is still unclear. This study aimed to evaluate the effect and mechanism of All-Trans Retinoic Acid (ATRA) on endothelial dysfunction induced by diabetes. METHODS: In the present study, streptozotocin (STZ)-induced diabetic rats and high glucose (HG)-induced human umbilical vein endothelial cells (HUVECs) were observed, and the effects of ATRA on HG-induced endothelial dysfunction and ferroptosis were evaluated. RESULTS: ATRA treatment improved impaired vasorelaxation in diabetic aortas in an endothelium-dependent manner, and this effect was accompanied by an increase in the NO concentration and eNOS expression. Ferroptosis, characterized by lipid peroxidation and iron overload induced by HG, was improved by ATRA administration, and a ferroptosis inhibitor (ferrostatin-1, Fer-1) improved endothelial function to a similar extent as ATRA. In addition, the inactivation of phosphoinositol-3-kinase (PI3K)/protein kinases B (AKT) and Yes-Associated Protein (YAP) nuclear localization induced by HG were reversed by ATRA administration. Vascular ring relaxation experiments showed that PI3K/AKT activation and YAP inhibition had similar effects on ferroptosis and endothelial function. However, the vasodilative effect of retinoic acid was affected by PI3K/AKT inhibition, and the inhibitory effects of ATRA on ferroptosis and the improvement of endothelial function were dependent on the retinoic acid receptor. CONCLUSION: ATRA could improve vascular endothelial dysfunction by inhibiting PI3K/AKT/YAP-mediated ferroptosis induced by HG, which provides a new idea for the treatment of vascular lesions in diabetes.

Neuroprotective effects of ellorarxine in neuronal models of degeneration.

Introduction: Retinoic acid (RA) was first recognised to be important for the central nervous system (CNS) in its developmental regulatory role and, given this action, it has been proposed in the adult CNS to regulate plasticity and promote regeneration. These types of roles have included support of neurogenesis, induction of neurite outgrowth, and protection from neuronal death. These functions are predominantly mediated by the retinoic acid receptor (RAR) transcription factor, and hence agonists for the RARs have been tested in a variety of models of neurodegeneration. This present study employs several in vitro models less explored for the action of RAR agonists to reverse neurodegeneration. Methods: A series of assays are used in which neuronal cells are placed under the types of stress that have been linked to neurodegeneration, in particular amyotrophic lateral sclerosis (ALS), and the neuroprotective influence of a new potent agonist for RAR, ellorarxine, is tested out. In these assays, neuronal cells were subjected to excitotoxic stress induced by glutamate, proteostasis disruption caused by epoxomicin, and oxidative stress leading to stress granule formation triggered by sodium arsenite. Results: Ellorarxine effectively reversed neuronal death in excitotoxic and proteostasis disruption assays and mitigated stress granule formation induced by sodium arsenite. This study also highlights for the first time the novel observation of RAR modulation of stress granules, although it is unknown whether this change in stress granules will be neuroprotective or potentially regenerative. Furthermore, the distribution of RAR agonists following intraperitoneal injection was assessed in mice, revealing preferential accumulation in the central nervous system, particularly in the spinal cord, compared to the liver. Gene expression studies in the spinal cord demonstrated that ellorarxine induces transcriptional changes at a low dose (0.01 mg/kg). Discussion: These findings underscore the therapeutic potential of RAR agonists, such as ellorarxine, for ALS and potentially other neurodegenerative diseases.

13-cis Retinoic Acid-Mediated Modulation of Human Meibomian Gland Epithelial Cells Development: Implications for In Vitro Modeling of Meibomian Gland Dysfunction.

Purpose: This study aimed to investigate the effect of 13-cis retinoic acid (13-cis RA) on human meibomian gland epithelial cells (HMGECs) and explore the potential of using this experimental model as an in vitro approach for studying meibomian gland dysfunction (MGD). Methods: First, HMGECs were cultured with 13-cis RA at different doses and times, and cell viability and proliferation rates were assessed to determine the appropriate stimulation concentration and time. Subsequently, during the proliferation stage, the expression of proliferation, inflammation, and oxidative stress genes and their products were evaluated. The meibum synthesis capacity was determined during the differentiation stage. Additionally, the peroxisome proliferator-activated receptor gamma (PPARγ) antagonist GW9662 was used as a control to assess the impact of 13-cis RA on PPARγ. Results: 13-cis RA significantly inhibited cell viability and proliferation in a time-dose response manner. Under the stimulation of 2 and 5 µM for 48 h during the proliferation stage, a significant decrease was observed in the expression of cell proliferation markers Ki67, antioxidant SOD-2, and Nrf-2. However, the expression of the pro-inflammatory factors IL-1ß, IL-8, MMP9, and oxidative stress markers NOX-4 and reactive oxygen species increased. During the differentiation stage, it suppressed meibum synthesis and the expression of meibocyte differentiation-related proteins adipose differentiation-associated protein 4 (ADFP4), elongation of very long chain fatty acid protein 4 (ELOVL4), sterol regulatory element-binding protein 2 (SREBP-2), and PPARγ. Conclusion: 13-cis RA inhibited cell viability, promoted inflammation and oxidative stress, and suppressed meibum synthesis through the PPARγ pathway. Our study shed light on the effect of 13-cis RA on HMGECs and provided a promising direction for studying MGD in vitro.

Retinoic acid-induced differentiation and oxidative stress inhibitors increase resistance of human neuroblastoma cells to La Crosse virus-induced cell death.

La Crosse Virus (LACV) encephalitis patients are at risk for long-term deficits in cognitive function due to neuronal apoptosis following virus infection. However, the specific etiology underlying neuronal damage remains elusive. In this study, we examined how differentiation and mitotic inhibition of neuroblastoma cells influence their susceptibility to LACV infection and cell death. Treatment of SH-SY5Y cells with retinoic acid induced a neuronal cell phenotype which was similarly susceptible to LACV infection as untreated cells but had significantly delayed virus-induced cell death. Protein and RNA transcript analysis showed that retinoic acid-treated cells had decreased oxidative stress responses to LACV infection compared to untreated cells. Modulation of oxidative stress in untreated cells with specific compounds also delayed cell death, without substantially impacting virus production. Thus, the oxidative stress response of neurons to virus infection may be a key component of neuronal susceptibility to virus-induced cell death. IMPORTANCE: Encephalitic viruses like La Crosse Virus (LACV) infect and kill neurons. Disease onset and progression is rapid meaning the time frame to treat patients before significant and long-lasting damage occurs is limited. Examining how neurons, the primary cells infected by LACV in the brain, resist virus-induced cell death can provide avenues for determining which pathways to target for effective treatments. In the current study, we studied how changing neuroblastoma growth and metabolism with retinoic acid treatment impacted their susceptibility to LACV-induced cell death. We utilized this information to test compounds for preventing death in these cells.

Single-cell RNA sequencing profiling of mouse cardiac cells in response to retinoic acid.

Congenital heart disease (CHD) is the leading cause of birth defect-related mortality. CHD is a multifactorial, complex disease involving environmental factors playing important roles. To elucidate the cardiac cellular and molecular mechanisms of cardiac malformation, we administered pregnant mice with a single dose of all-trans retinoic acid (RA) at E8.5, as the CHD model. We performed single-cell RNA sequencing on cardiac cells from developing mouse hearts spanning from E8.5 to E17.5 after RA administration. A total of 69,447 cells were obtained from seven developmental stages ranging from E8.5 to E17.5. RA significantly impacted various CM subpopulations, particularly the outflow tract CMs at E9.0 by reduction of Tdgf1 expression. RA also influences the transition of endocardial-to-mesenchymal cells by decreasing the Stmn2 levels, which may contribute to abnormal valve development. In addition, RA altered the metabolic pattern of epicardial cells at E11.5 and promoted its differentiation potential. Taken together, these results are valuable for the development of preventive and therapeutic strategies for CHDs.

Dual and opposing role of retinoic acid receptor signaling in mesenchymal stem cells for tendon ossification in mice.

Heterotopic ossification is abnormal bone formation in soft tissues that occurs primarily after injury and major surgery. This condition often causes local pain and limits joint motion in the affected limb. Currently, there is no effective treatment or prophylaxis for this condition other than surgical removal of the lesion. Recent studies suggest that retinoic acid receptor (RAR) agonists are effective in suppressing heterotopic ossification in patients with Fibrodysplasia Ossificans Progressiva, a congenital disorder characterized by progressive ossification of soft tissue, by suppressing the aberrant differentiation of mesenchymal stem cells in muscle. In this study, we aimed to elucidate the potential use of RAR agonists in suppressing injury-induced ectopic tendon ossification using a mouse Achilles tenotomy model. Contrary to our initial hypothesis, administration of RAR agonists throughout the experimental period (5 weeks) accelerated ectopic tendon ossification in our model. Of note, in vitro differentiation experiments using tendon-derived mesenchymal stem cells revealed that RAR agonists play opposing roles in osteogenic and chondrogenic differentiation, promoting the former and suppressing the latter. Indeed, we found that RAR agonists suppressed tendon ossification when administered before cartilage nodule formation, but promoted it when administered after. These results suggest that RAR agonists have a dual and opposing effect on tendon ectopic ossification, depending on the duration and timing of their administration. Our data may provide a basis for further investigation of the potential use of RAR agonists in the treatment of injury-induced heterotopic ossification.

Characterization of the C17.2 cell line as testing system for endocrine disruption-induced developmental neurotoxicity.

Hormone signaling plays an essential role during fetal life and is vital for brain development. Endocrine-disrupting chemicals can interfere with the hormonal milieu during this critical time-period, disrupting key neurodevelopmental processes. Hence, there is a need for the development of assays that evaluate developmental neurotoxicity (DNT) induced by an endocrine mode of action. Herein, we evaluated the applicability of the neural progenitor C17. 2 cell-line, as an in vitro test system to aid in the detection of endocrine disruption (ED) induced DNT. For this, C17.2 cells were exposed during 10 days of differentiation to agonists and antagonists of the thyroid hormone (Thr), glucocorticoid (Gr), retinoic acid (Rar), retinoic x (Rxr), oxysterols (Lxr), estrogen (Er), androgen (Ar), and peroxisome proliferator activated delta (Pparß/δ) receptors, as well as to the agonist of the vitamin D (Vdr) receptor. Upon exposure and differentiation, neuronal morphology (neurite outgrowth and branching), and the percentage of neurons in culture were assessed by immunofluorescence. For this, the cells were incubated with Hoechst (nuclear staining) and stained for ßIII-tubulin (neuronal marker). The C17.2 cells were responsive to the Rar, Rxr and Pparß/δ agonists which decreased neurite outgrowth and branching. Additionally, exposure to the Gr agonist increased the number of cells differentiating into neurons, while exposure to the Rxr agonist had the opposite effect. With this approach, we have identified that the C17.2 cells are responsive to Gr, Rar, Rxr, and Pparß/δ agonists, hence contributing to the development of test systems for hazard assessment of ED-induced DNT.

Endocrine disrupting chemicals (EDCs) interfere with hormonal signaling. As hormones play a vital role for an organism's development, EDC exposure is of high concern. In European regulations, the use of a chemical can be restricted if its toxicity is mediated by hormonal interference. A number of EDCs affect brain development. However, in animal tests, it is impossible to prove that a chemical induces developmental neurotoxicity (DNT) via endocrine disruption (ED). Furthermore, the regulatory DNT tests require large amounts of animals. Thus, there is an urgent need for in vitro test systems to identify ED-induced DNT. Herein we present the development of such a method based on the murine neural progenitor cell-line C17.2 with which neuronal differentiation processes can be mimicked. We show that differentiation of C17.2 cells are sensitive to retinoid, glucocorticoid, and peroxisome proliferator activated receptor signaling disruption, thus providing an alternative method for identifying ED-induced DNT.

Unveiling the role of RARs in stomach adenocarcinoma: clinical implications and prognostic biomarkers.

Background: Retinoic acid receptors (RARs) family are known to play a significant role in the occurrence and development of tumors. However, the relationship between RARs and stomach adenocarcinoma (STAD) has not yet been clearly identified. The aim of this study is to evaluate the expression profile and clinical value of the RARs family in STAD. Methods: The expression level, clinical characteristics, prognostic value, immunity-related evaluations, genetic alteration and methylation site of RARs in STAD were explored using a series of online databases including gene expression profiling interactive analysis (GEPIA), tumor immune estimation resource (TIMER), University of Alabama at Birmingham cancer data (UALCAN), Human Protein Atlas (HPA), Kaplan-Meier plotter, gene set cancer analysis (GSCA), cBioPortal, MethSurv, GeneMANIA, LinkedOmics, Metascape, Search tool for the retrieval of interacting genes (STRING), tumor immune single-cell hub (TISCH) and cancer cell line encyclopedia (CCLE). Results: We discovered dramatically increased expression of RARA and decreased expression of RARB in STAD tissues, and many clinical variables were closely related to RARs. Notably, higher expressions of RARA and RARB as well as lower expression of RARG correlated with worse overall survival (OS) for STAD patients. The clinical value of prognostic model indicated that RARs were identified to be potential prognostic biomarkers for STAD patients. Moreover, RARB was closely related to immune cell infiltration, which had effect on the role of RARB in STAD prognosis. And the genetic alteration of RARB was significantly associated with the longer disease-free survival (DFS) of STAD patients. Additionally, some CpG sites of the RARs family were related with the prognosis of STAD patients. Functional enrichment analyses indicated that several pathways in STAD might be pivotal pathways regulated by RARs. At the single-cell level, there was some extent of infiltration of tumor microenvironment-related cells in the RARs expression in STAD. Conclusions: Our results evaluated the expression profile and clinical values of RARs in patients with STAD, which provided a basis for future in-depth exploration of the specific mechanisms of each member of RARs in STAD.

Visual Hallucinations With Arsenic Trioxide Therapy in Acute Promyelocytic Leukemia.

A 68-year-old male with a history of diabetes and hypertension was diagnosed with acute promyelocytic leukemia (APML). He underwent induction therapy with all-trans retinoic acid (ATRA) and arsenic trioxide. He had a complete hematologic response and was initiated on consolidation therapy with arsenic trioxide (0.15 mg/kg/day intravenous (IV)) and ATRA (45 mg/per meter square of body surface area/day IV). He developed blurred vision and floaters after a few days. Soon after, he felt that his diabetic neuropathy had suddenly worsened. The floaters and flashing lights worsened and morphed into visual hallucinations. He reported seeing figures watching him from the corner of the room. He was admitted and head imaging was unremarkable. Routine labs did not show anything unusual. Arsenic trioxide therapy was held. The hallucinations gradually started decreasing and eventually subsided after around eight weeks. ATRA was continued but arsenic was permanently discontinued. Arsenic is known to cause poisoning if exposed in significant amounts. The arsenic dose used for APML is substantially low (0.15 mg/kg/day IV). We delineate this unanticipated case of arsenic toxicity leading to severe neurological symptoms like visual hallucinations which has not been previously reported in the literature. It is imperative to closely monitor patients who are on arsenic therapy and inform them about possible rare toxicities.

Leucocytosis during all-trans retinoic acid and arsenic trioxide treatment in acute promyelocytic leukaemia.

In patients with acute promyelocytic leukaemia (APL), differentiation syndrome (DS) is a life-threatening complication caused by the differentiating effect of all-trans retinoic acid (ATRA) and/or arsenic trioxide (ATO). Leucocytosis is frequently observed during induction therapy for APL and is intimately associated with the development of DS and its severity. The management of DS is particularly important due to the high likelihood of excellent outcomes for APL patients who successfully complete induction therapy. Commentary on: Cicconi et al. Leukocytosis during induction therapy with all-trans-retinoic acid and arsenic trioxide in acute promyelocytic leukemia predicts for differentiation syndrome and treatment-related complications. Br J Haematol 2024 (Online ahead of print). doi: 10.1111/bjh.19759.

Exosomal miR-205-5p contributes to the immune liver injury induced by trichloroethylene: Pivotal role of RORα mediating M1 Kupffer cell polarization.

Trichloroethylene (TCE) is a common environmental contaminant that can induce occupational dermatitis medicamentosa-like TCE (ODMLT), where the liver damage is the most common complication. The study aims to uncover the underlying mechanism of TCE-sensitization-induced liver damage by targeting specific exosomal microRNAs (miRNAs). Among the enriched serum exosomal miRNAs of ODMLT patients, miR-205-5p had a significant correlation coefficient with the liver function damage indicators. Moreover, retinoic acid receptor-related orphan receptor α (RORα) was identified as a direct target of miR-205-5p via specific binding. Further experiments showed that kupffer cells (KCs) underwent M1 phenotypic and functional changes in liver injury induced by TCE which were alleviated by reducing the expression of miR-205-5p. However, this alleviation was reversed by the RORα antagonist SR1001. In vitro experiments showed that miR-205-5p promoted M1 polarization of macrophages and enhanced the secretion of inflammatory factors by regulating RORα. An increase in RORα reversed the polarization direction of M1-type macrophages and reduced the secretion of proinflammatory factors. In addition, pretreatment of mice with SR1078, a specific RORα agonist, effectively blocked M1 polarization of KCs and reduced the severity of TCE-induced liver injury. Our study uncovers that miR-205-5p regulates KC M1 polarization by targeting RORα in immune liver injury induced by TCE sensitization, providing new insight into the molecular mechanisms and new therapeutic targets for ODMLT.

Targeting Androgen, Thyroid Hormone, and Vitamin A and D Receptors to Treat Prostate Cancer.

The nuclear hormone family of receptors regulates gene expression. The androgen receptor (AR), upon ligand binding and homodimerization, shuttles from the cytosol into the nucleus to activate gene expression. Thyroid hormone receptors (TRs), retinoic acid receptors (RARs), and the vitamin D receptor (VDR) are present in the nucleus bound to chromatin as a heterodimer with the retinoid X receptors (RXRs) and repress gene expression. Ligand binding leads to transcription activation. The hormonal ligands for these receptors play crucial roles to ensure the proper conduct of very many tissues and exert effects on prostate cancer (PCa) cells. Androgens support PCa proliferation and androgen deprivation alone or with chemotherapy is the standard therapy for PCa. RARγ activation and 3,5,3'-triiodo-L-thyronine (T3) stimulation of TRß support the growth of PCa cells. Ligand stimulation of VDR drives growth arrest, differentiation, and apoptosis of PCa cells. Often these receptors are explored as separate avenues to find treatments for PCa and other cancers. However, there is accumulating evidence to support receptor interactions and crosstalk of regulatory events whereby a better understanding might lead to new combinatorial treatments.

The order of green and red LEDs irradiation affects the neural differentiation of human umbilical cord matrix-derived mesenchymal cells.

Different wavelengths emitted from light-emitting diodes (LEDs) are known as an influential factor in proliferation and differentiation of various cell types. Since human umbilical cord matrix-derived mesenchymal cells (hUCMs) are ideal tools for human regenerative medicine clinical trials and stem cell researches, in the present study we investigated the neurogenesis effects of single and intermittent green and red LED irradiation on hUCM cells. Exposure of hUCMs to single and intermittent green (530 nm, 1.59 J/cm2) and red (630 nm, 0.318 J/cm2) lights significantly increased the expression of specific genes including nestin, ß-tubulin III and Olig2. Additionally, immunocytochemical analysis confirmed the expression of specific neural-related proteins including nestin, ß-tubulin III, Olig2 and GFAP. Also, alternating exposure of hUCM cells to green and red lights increased the expression of some neural markers more than either light alone. Further research are required to develop the application of LED irradiation as a useful tool for therapeutic purposes including neural repair and regeneration.

C286, an orally available retinoic acid receptor ß agonist drug, regulates multiple pathways to achieve spinal cord injury repair.

Retinoic acid receptor ß2 (RARß2) is an emerging therapeutic target for spinal cord injuries (SCIs) with a unique multimodal regenerative effect. We have developed a first-in-class RARß agonist drug, C286, that modulates neuron-glial pathways to induce functional recovery in a rodent model of sensory root avulsion. Here, using genome-wide and pathway enrichment analysis of avulsed rats' spinal cords, we show that C286 also influences the extracellular milieu (ECM). Protein expression studies showed that C286 upregulates tenascin-C, integrin-α9, and osteopontin in the injured cord. Similarly, C286 remodulates these ECM molecules, hampers inflammation and prevents tissue loss in a rodent model of spinal cord contusion C286. We further demonstrate C286's efficacy in human iPSC-derived neurons, with treatment resulting in a significant increase in neurite outgrowth. Additionally, we identify a putative efficacy biomarker, S100B, which plasma levels correlated with axonal regeneration in nerve-injured rats. We also found that other clinically available retinoids, that are not RARß specific agonists, did not lead to functional recovery in avulsed rats, demonstrating the requirement for RARß specific pathways in regeneration. In a Phase 1 trial, the single ascending dose (SAD) cohorts showed increases in expression of RARß2 in white blood cells correlative to increased doses and at the highest dose administered, the pharmacokinetics were similar to the rat proof of concept (POC) studies. Collectively, our data suggests that C286 signalling in neurite/axonal outgrowth is conserved between species and across nerve injuries. This warrants further clinical testing of C286 to ascertain POC in a broad spectrum of neurodegenerative conditions.

Therapy-related myeloid neoplasms following curative treatment of acute promyelocytic leukemia: incidence, correlation with therapeutic regimen, and future directions.

All-trans retinoic acid (ATRA) and arsenic trioxide (ATO) have revolutionized the treatment of acute promyelocytic leukemia (APL), offering a cure rate of > 80%. Along with improved survival, the long-term consequences of anti-APL therapy are becoming increasingly apparent, including potential therapy-related myeloid neoplasms (t-MNs). T-MNs are well known to arise after cytotoxic chemotherapy, but the leukemogenic potential of regimens utilizing only ATRA/ATO is not well established. The objective of this study is to examine the incidence, long-term risk, and clinicopathologic features of t-MNs arising after anti-APL therapy and how they correlates with the therapeutic regimen employed. We retrospectively collected treated APL patients between 01/2001 and 02/2021, categorized them into ATRA/ATO + chemo and ATRA/ATO groups based on the regimen used, and evaluated for the development of t-MN. A total of 110 APL patients were identified, including 67 (61%) treated with ATRA/ATO + chemo and 43 (39%) treated with ATRA/ATO only. Overall, 4/110 (3.6%) patients developed t-MNs, with all four emerging in the ATRA/ATO + chemo group. Ultimately, the incidence of t-MN in ATRA/ATO + chemo group was significantly higher compared with ATRA/ATO only group(5.97% vs. 0.0%, respectively; p = 0.0289). Our data spanning over two decades suggests that conventional chemotherapy for APL is associated with a small but significant risk of t-MN, whereas ATR/ATO does not carry this risk. This takes on new significance, considering several recent and ongoing trials have shown that a chemotherapy-free approach might become feasible for all risk APL types in the near future. Consequently, the omission of leukemogenic and arguably unnecessary chemotherapy from APL regimens may reduce the incidence of t-MNs in long-term survivors without sacrificing their cure rates.

The potential of retinoic acid receptors as prognostic biomarkers and therapeutic targets in gastric cancer.

Background: Gastric cancer is a heterogeneous collection of tumors characterized by low survival rates. All-trans retinoic acid (retinoic-acid) is a clinically useful therapeutic agent belonging to the chemical family of retinoids, which consists of both natural and synthetic derivatives of vitamin-A. Retinoids are essential components of the normal diet and they regulate different physiological processes. From a therapeutic point of view, retinoic-acid is the first example of clinically useful differentiating agent. Indeed, the differentiating properties of this compound have promoted the use of retinoic-acid as a standard of care in Acute-Promyelocytic-Leukemia, a rare form of acute myeloid leukemia. In this study, we determine the RNA expression of the six isoforms of Retinoic-Acid-Receptors (RARα/RARß/RARγ/RXRα/RXRß/RXRγ) in view of their potential use as gastric cancer progression markers and/or therapeutic targets. In addition, we evaluate associations between the expression of these receptors and a simplified molecular classification of stomach tumors as well as the clinical characteristics of the cohort of patients analyzed. Finally, we define the prognostic value of the various Retinoic-Acid-Receptors in gastric cancer. Methods: In this single institution and retrospective RAR-GASTRIC study, we consider 55 consecutive gastric cancer patients. We extract total RNA from the pathological specimens and we perform a NanoString Assay using a customized panel of genes. This allows us to determine the expression levels of the RAR and RXR mRNAs as well as other transcripts of interest. Results: Our data demonstrate ubiquitous expression of the RAR and RXR mRNAs in gastric cancers. High levels of RARα, RARß, RXRα and RXRß show a significant association with stage IV tumors, "de novo" metastatic disease, microsatellite-stable-status, epithelial-to-mesenchymal-transition, as well as PIK3CA and TP53 expression. Finally, we observe a worse overall-survival in gastric cancer patients characterized by high RARα/RARß/RARγ/RXRß mRNA levels. Conclusions: In gastric cancer, high expression levels of RARα/RARß/RARγ/RXRß transcripts are associated with poor clinical and molecular characteristics as well as with reduced overall-survival. Our data are consistent with the idea that RARα, RARß, RARγ and RXRß represent potential prognostic markers and therapeutic targets of gastric cancer.

Chemical Derivatization and Paper Spray Ionization Mass Spectrometry for Fast Screening of Retinoic Acid in Cosmetics.

As a prescription drug, retinoic acid is listed as a banned cosmetic additive in the EU and China regulations. Currently, spectrophotometric methods, including thin-layer chromatography (TLC), high-performance liquid chromatography (HPLC), and HPLC-MS/MS, are commonly used for the determination of retinoic acid. As these conventional methods require complex pretreatment and are time-consuming, chemical derivatization combined with paper spray ionization mass spectrometry was developed for the fast detection of retinoic acid in cosmetics. N,N-dimethylpiperazine iodide (DMPI) was utilized as a derivatization reagent. Carboxylic acid in retinoic acid was derivatized to carry a positive charge and was subjected to mass spectrometry analysis. Results showed that compared with non-derivatized compounds, the detection limit was increased by about 50 times. The linearity in the range of 0.005-1 µg·mL-1 was good. The limit of detection (LOD) was 0.0013 µg·mL-1, and the limit of quantification (LOQ) was 0.0043 µg·mL-1. The recoveries of spiked samples were in the range of 95-105%, and the RSDs were below 5%. Derivatization and paper spray ionization MS render a quick, sensitive, and accurate method for the detection of retinoic acid in a complex matrix.

Modeling Epithelial Homeostasis and Perturbation in Three-Dimensional Human Esophageal Organoids.

Background: Esophageal organoids from a variety of pathologies including cancer are grown in Advanced Dulbecco's Modified Eagle Medium-Nutrient Mixture F12 (hereafter ADF). However, the currently available ADF-based formulations are suboptimal for normal human esophageal organoids, limiting the ability to compare normal esophageal organoids with those representing a given disease state. Methods: We have utilized immortalized normal human esophageal epithelial cell (keratinocyte) lines EPC1 and EPC2 and endoscopic normal esophageal biopsies to generate three-dimensional (3D) organoids. To optimize the ADF-based medium, we evaluated the requirement of exogenous epidermal growth factor (EGF) and inhibition of transforming growth factor-(TGF)-ß receptor-mediated signaling, both key regulators of the proliferation of human esophageal keratinocytes. We have modeled human esophageal epithelial pathology by stimulating esophageal 3D organoids with interleukin (IL)-13, an inflammatory cytokine, or UAB30, a novel pharmacological activator of retinoic acid signaling. Results: The formation of normal human esophageal 3D organoids was limited by excessive EGF and intrinsic TGFß-receptor-mediated signaling. Optimized HOME0 improved normal human esophageal organoid formation. In the HOME0-grown organoids, IL-13 and UAB30 induced epithelial changes reminiscent of basal cell hyperplasia, a common histopathologic feature in broad esophageal disease conditions including eosinophilic esophagitis. Conclusions: HOME0 allows modeling of the homeostatic differentiation gradient and perturbation of the human esophageal epithelium while permitting a comparison of organoids from mice and other organs grown in ADF-based media.