Characterization of a model of liver regeneration: Role of hedgehog signaling in experimental hepatic amoebiasis.

The development of amoebic liver abscess (ALA) leads to liver necrosis, accompanied by an exacerbated inflammatory response and the formation of multiple granulomas. Adequate management of the infection through the administration of treatment and the timely response of the organ to the damage allows the injury to heal with optimal regeneration without leaving scar tissue, which does not occur in other types of damage such as viral hepatitis that may conducts to fibrosis or cirrhosis. The Hedgehog signaling pathway (Hh) is crucial in the embryonic stage, while in adults it is usually reactivated in response to acute or chronic injuries, regeneration, and wound healing. In this work, we characterized Hh in experimental hepatic amoebiasis model, with the administration of treatment with metronidazole, as well as a pathway inhibitor (cyclopamine), through histological and immunohistochemical analyses including an ultrastructure analysis through transmission electron microscopy. The results showed an increase in the percentage of lesions obtained, a decrease in the presence of newly formed hepatocytes, a generalized inflammatory response, irregular distribution of type I collagen accompanied by the presence of fibroblast-type cells and a decrease in effector cells of this pathway. These results constitute the first evidence of the association of the activation of Hh with the liver regeneration process in experimental amebiasis.

The Primary Cilium and its Hedgehog Signaling in Nociceptors Contribute to Inflammatory and Neuropathic Pain.

The primary cilium, a 1-3 µm long hair-like structure protruding from the surface of almost all cells in the vertebrate body, is critical for neuronal development and also functions in the adult. As the migratory neural crest settles into dorsal root ganglia (DRG) sensory neurons elaborate a single primary cilium at their soma that is maintained into adult stages. While it is not known if primary cilia are expressed in nociceptors, or their potential function in the mature DRG neuron, recent studies have shown a role for Hedgehog, whose signaling demonstrates a dependence on primary cilia, in nociceptor sensitization. Here we report the expression of primary cilia in rat and mouse nociceptors, where they modulate mechanical nociceptive threshold, and contribute to inflammatory and neuropathic pain. When siRNA targeting Ift88, a primary cilium-specific intraflagellar transport (IFT) protein required for ciliary integrity, was administered by intrathecal injection, in the rat, it resulted in loss of Ift88 mRNA in DRG, and primary cilia in neuronal cell bodies, which was associated with an increase in mechanical nociceptive threshold, and abrogation of hyperalgesia induced by the pronociceptive inflammatory mediator, prostaglandin E2, and painful peripheral neuropathy induced by a neurotoxic chemotherapy drug, paclitaxel. To provide further support for the role of the primary cilium in nociceptor function we also administered siRNA for another IFT protein, Ift52. Ift52 siRNA results in loss of Ift52 in DRG and abrogates paclitaxel-induced painful peripheral neuropathy. Attenuation of Hedgehog-induced hyperalgesia by Ift88 knockdown supports a role for the primary cilium in the hyperalgesia induced by Hedgehog, and attenuation of paclitaxel chemotherapy-induced neuropathy (CIPN) by cyclopamine, which attenuates Hedgehog signaling, suggests a role of Hedgehog in CIPN. Our findings support a role of nociceptor primary cilia in the control of mechanical nociceptive threshold and in inflammatory and neuropathic pain, the latter, at least in part, Hedgehog dependent.

Pharmacology of Veratrum californicum Alkaloids as Hedgehog Pathway Antagonists.

Veratrum californicum contains steroidal alkaloids that function as inhibitors of hedgehog (Hh) signaling, a pathway involved in the growth and differentiation of cells and normal tissue development. This same Hh pathway is abnormally active for cell proliferation in more than 20 types of cancer. In this current study, alkaloids have been extracted from the root and rhizome of V. californicum, followed by their separation into five fractions using high performance liquid chromatography. Mass spectrometry was used to identify the presence of twenty-five alkaloids, nine more than are commonly cited in literature reports, and the Bruker Compass Data Analysis software was used to predict the molecular formula for every detected alkaloid. The Gli activity of the raw extract and each fraction were compared to 0.1 µM cyclopamine, and fractions 1, 2, and 4 showed increased bioactivity through suppression of the Hh signaling pathway. Fractions 2 and 4 had enhanced bioactivity, but fraction 1 was most effective in inhibiting Hh signaling. The composition of fraction 1 consisted of veratrosine, cycloposine, and potential isomers of each.

Increasing Ciliary ARL13B Expression Drives Active and Inhibitor-Resistant Smoothened and GLI into Glioma Primary Cilia.

ADP-ribosylation factor-like protein 13B (ARL13B), a regulatory GTPase and guanine exchange factor (GEF), enriches in primary cilia and promotes tumorigenesis in part by regulating Smoothened (SMO), GLI, and Sonic Hedgehog (SHH) signaling. Gliomas with increased ARL13B, SMO, and GLI2 expression are more aggressive, but the relationship to cilia is unclear. Previous studies have showed that increasing ARL13B in glioblastoma cells promoted ciliary SMO accumulation, independent of exogenous SHH addition. Here, we show that SMO accumulation is due to increased ciliary, but not extraciliary, ARL13B. Increasing ARL13B expression promotes the accumulation of both activated SMO and GLI2 in glioma cilia. ARL13B-driven increases in ciliary SMO and GLI2 are resistant to SMO inhibitors, GDC-0449, and cyclopamine. Surprisingly, ARL13B-induced changes in ciliary SMO/GLI2 did not correlate with canonical changes in downstream SHH pathway genes. However, glioma cell lines whose cilia overexpress WT but not guanine exchange factor-deficient ARL13B, display reduced INPP5e, a ciliary membrane component whose depletion may favor SMO/GLI2 enrichment. Glioma cells overexpressing ARL13B also display reduced ciliary intraflagellar transport 88 (IFT88), suggesting that altered retrograde transport could further promote SMO/GLI accumulation. Collectively, our data suggest that factors increasing ARL13B expression in glioma cells may promote both changes in ciliary membrane characteristics and IFT proteins, leading to the accumulation of drug-resistant SMO and GLI. The downstream targets and consequences of these ciliary changes require further investigation.

Hardening of Respiratory Syncytial Virus Inclusion Bodies by Cyclopamine Proceeds through Perturbation of the Interactions of the M2-1 Protein with RNA and the P Protein.

Respiratory syncytial virus (RSV) RNA synthesis takes place in cytoplasmic viral factories also called inclusion bodies (IBs), which are membrane-less organelles concentrating the viral RNA polymerase complex. The assembly of IBs is driven by liquid-liquid phase separation promoted by interactions between the viral nucleoprotein N and the phosphoprotein P. We recently demonstrated that cyclopamine (CPM) inhibits RSV multiplication by disorganizing and hardening IBs. Although a single mutation in the viral transcription factor M2-1 induced resistance to CPM, the mechanism of action of CPM still remains to be characterized. Here, using FRAP experiments on reconstituted pseudo-IBs both in cellula and in vitro, we first demonstrated that CPM activity depends on the presence of M2-1 together with N and P. We showed that CPM impairs the competition between P and RNA binding to M2-1. As mutations on both P and M2-1 induced resistance against CPM activity, we suggest that CPM may affect the dynamics of the M2-1-P interaction, thereby affecting the relative mobility of the proteins contained in RSV IBs. Overall, our results reveal that stabilizing viral protein-protein interactions is an attractive new antiviral approach. They pave the way for the rational chemical optimization of new specific anti-RSV molecules.

Hedgehog signaling is essential in the regulation of limb regeneration in the Chinese mitten crab, Eriocheir sinensis.

Tissue autotomy is a unique adaptive response to environmental stress, followed by regeneration process compensating for the loss of body parts. The crustaceans present remarkable activity of appendage autotomy and regeneration, however, the molecular mechanism is still unclear. In this study, the Eriocheir sinensis Hedgehog (EsHH) and Smoothened (EsSMO) were identified in the regenerative limbs, and the function of Hedgehog signaling pathway on limb regeneration was evaluated. At the blastema growth stage of limb regeneration, the expression of EsHH and EsSMO was up-regulated in response to limb autotomy stress, and down-regulated at blastema differentiation stage. To clarify the effect of Hedgehog pathway during limb regeneration, the regenerative efficiency was evaluated with Smoothened inhibitor cyclopamine or RNAi (ds-HH) injection. We observed that the regenerative efficiency was significantly repressed with blockage of Hedgehog pathway at both the basal growth stage and the proecdysial growth stage, which was indicated by the delay of wound healing and blastema growth, as well as a decrease in the size of newly formed limbs. In addition, gene expression and BrdU incorporation assay showed that the proliferation and myogenic differentiation of blastema cells were suppressed with either cyclopamine or ds-HH injection. Thus, these results suggest that Hedgehog signaling pathway is essential for the establishment of limb regeneration in E. sinensis through promoting the proliferation and myogenic differentiation of blastema cells.

Screening antivirals with a mCherry-expressing recombinant bovine respiratory syncytial virus: a proof of concept using cyclopamine.

Bovine respiratory syncytial virus (BRSV) is a pathogenic pneumovirus and a major cause of acute respiratory infections in calves. Although different vaccines are available against BRSV, their efficiency remains limited, and no efficient and large-scale treatment exists. Here, we developed a new reverse genetics system for BRSV expressing the red fluorescent protein mCherry, based on a field strain isolated from a sick calf in Sweden. Although this recombinant fluorescent virus replicated slightly less efficiently compared to the wild type virus, both viruses were shown to be sensitive to the natural steroidal alkaloid cyclopamine, which was previously shown to inhibit human RSV replication. Our data thus point to the potential of this recombinant fluorescent BRSV as a powerful tool in preclinical drug discovery to enable high throughput compound screening.

Cyclopamine targeting hedgehog modulates nuclear control of the osteoblast activity.

It is known that cellular events underlying the processes of bone maintenance, remodeling, and repair have their basis in the embryonic production of bone. Shh signaling is widely described developing important morphogenetic control in bone by modifying the activity of osteoblast. Furthermore, identifying whether it is associated with the modulation of nuclear control is very important to be the basis for further applications. Experimentally, osteoblasts were exposed with cyclopamine (CICLOP) considering up to 1 day and 7 days, here considered an acute and chronic responses respectively. Firstly, we have validated the osteogenic model in vitro by exposing the osteoblasts to classical differentiating solution up to 7 days to allow the analysis of alkaline phosphatase and mineralization. Conversely, our data shows that differentiating osteoblasts present higher activity of inflammasome-related genes, while Shh signaling members were lower, suggesting a negative feedback between them. Thereafter, to better know about the role of Shh signaling on this manner, functional assays using CICLOP (5 µM) were performed and the data validates the previously hypothesis that Shh represses inflammasome related genes activities. Altogether, our data supports the anti-inflammatory effect of Shh signaling by suppressing Tnfα, Tgfß and inflammasome related genes during osteoblast differentiation, and this comprehension might support the understanding the molecular and cellular mechanisms related in bone regeneration by reporting molecular-related osteoblast differentiation.

Hydroxylated cyclopamine analogues from Veratrum californicum and their hedgehog pathway inhibiting activity.

Cyclopamine (1), the teratogenic steroidal alkaloid isolated from corn lily (Veratrum californicum), has recently gained renewed interest due to its anticancer potential, that has been translated into the FDA approval of three Hedgehog (Hh) pathway inhibiting antitumor drugs. A chemical analysis of mother liquors obtained from crystallization of cyclopamine, extracted from roots and rhizomes of V. californicum, resulted in the isolation of two unprecedented cyclopamine analogues, 18-hydroxycyclopamine (2) and 24R-hydroxycyclopamine (3), the first compounds of this class to show modifications on rings D-F. The stereostructures of these new natural compounds have been established based on a detailed MS and 1D/2D NMR investigation. The isolated compounds were evaluated with the dual-luciferase bioassay for their inhibition of the hedgehog pathway in comparison to cyclopamine, providing new insights into the structure-activity relationships for this class of compounds.

[Retracted] Hedgehog signaling is involved in the BMP9­induced osteogenic differentiation of mesenchymal stem cells.

Subsequently to the publication of the above paper, a concerned reader drew to the authors' attention that there were a number of overlapping data panels featured in the cellular images shown in Fig. 2C on p. 1644, and Figs. 3D and 4 on p. 1645, such that data that were allegedly obtained under different experimental conditions appeared to have been derived from some of the same original sources. Given the number of errors that had been made during the compilation of the figures in this article, the Editor of International Journal of Molecular Medicine has decided that this article should be retracted from the publication owing to a lack of overall confidence in the presented data. The authors were asked for an explanation to account for these concerns, but the Editorial Office did not receive a reply. The Editor apologizes to the readership for any inconvenience that might result from the retraction of this article. [International Journal of Molecular Medicine 35: 1641­1650, 2015; DOI: 10.3892/ijmm.2015.2172].

Veratrum parviflorum poisoning: identification of steroidal alkaloids in patient blood and breast milk.

INTRODUCTION: The Veratrum genus is composed of plants containing a diverse set of steroidal alkaloids. Veratrum plant material has been utilized for centuries as herbal medicines, however the alkaloids have such a low therapeutic index that they are not used in modern medicine. Here we report an incident of inadvertent ingestion of V. parviflorum by hikers in Georgia that allowed detection, and in several instances identification of alkaloids from the plant, and correlated their presence within patient blood and breast milk specimens. CASE HISTORY: Eight patients, three male and five female, presented in the spring of 2020 and 2021 with symptoms requiring emergent medical attention after ingestion of Veratrum parviflorum. All patients believed the plants to be a local native species of wild leek, Allium tricoccum, locally known as ramps. Plants were identified using photographs as well as fresh and cooked plant material provided by patients, in consultation with botanists at the University of Georgia Herbarium. Written consent was obtained from all patients for collection of blood and breast milk specimens for laboratory identification of Veratrum alkaloids. METHODS: V. parviflorum plant material, and patient serum and breast milk were analyzed by high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (HPLC-QTOF) to identify steroidal alkaloids. RESULTS: The V. parviflorum extract was confirmed to contain cyclopamine, veratramine, jervine, and muldamine. Two out of the eight patients had detectable concentrations of Veratrum alkaloids. Of the alkaloids identified in the plant, cyclopamine and jervine were detected within patient serum, and cyclopamine and veratramine were observed to be present in breast milk. DISCUSSION: Toxicity resulting from Veratrum steroidal alkaloids has primarily been reported from V. album and V. viride. This is the second report of V. parviflorum poisoning. The present work reports for the first time the presence of muldamine and jervine within V. parviflorum. This work provides the first instance of identification of Veratrum alkaloids in breast milk. Thus, the findings presented herein add to literature record causative agents contributing to the toxicity of V. parviflorum when ingested and potential for secondary poisoning through breastfeeding. CONCLUSION: V. parviflorum toxicity was observed to cause nausea, vomiting, hypotension, bradycardia, abdominal pain, light-headedness, blurred vision, and tingling in the arms. Patients experiencing mild symptoms improved with supportive care, IV fluids, and antiemetics, but hemodynamically unstable patients required atropine and vasopressors. This study demonstrated that more lipophilic Veratrum alkaloids can be passed along in breast milk, which suggests additional precautions may be critical to limit further poisonings.

Dhh signaling pathway regulates reconstruction of seminiferous tubule-like structure.

The reconstruction of a tubule-like structure in vitro has provided a promising system to analyze factors that drive morphogenesis and the underlying mechanisms. In this study, we took advantage of the inhibitor cyclopamine and a smoothened agonist to detect the role of the Dhh signaling pathway in the reconstructed tubule-like structure. Sertoli cells did not show polarity, rounded peritubular myoid cells and scattered Leydig cells were observed, combined with less laminin and lower proliferation of Leydig, peritubular myoid, germ, and Sertoli cells. However, in the presence of SAG, elongated peritubular myoid cells gathered at the bottom of polarized Sertoli cells, and most of the Leydig cells gathered at the outer part of the elongated peritubular myoid cells. Moreover, SAG promoted the secretion of laminin, assisting in the formation of the basal membrane and promoting the proliferation of Leydig, peritubular myoid, and germ cells. The level of Gli1 was significantly downregulated when treated with cyclopamine, whereas it was significantly upregulated when treated with SAG. These results indicate that the Dhh signaling pathway regulates the reconstruction of tubule-like structures by regulating the expression of Gli1.

Veratrum parviflorum: An Underexplored Source for Bioactive Steroidal Alkaloids.

Plants of the Veratrum genus have been used throughout history for their emetic properties, rheumatism, and for the treatment of high blood pressure. However, inadvertent consumption of these plants, which resemble wild ramps, induces life-threatening side effects attributable to an abundance of steroidal alkaloids. Several of the steroidal alkaloids from Veratrum spp. have been investigated for their ability to antagonize the Hedgehog (Hh) signaling pathway, a key pathway for embryonic development and cell proliferation. Uncontrolled activation of this pathway is linked to the development of various cancers; most notably, basal cell carcinoma and acute myeloid leukemia. Additional investigation of Veratrum spp. may lead to the identification of novel alkaloids with the potential to serve as chemotherapeutics. V. parviflorum is a relatively uncommon species of Veratrum that resides in the southeastern regions of North America. The phytochemical profile of this plant remains largely unexplored; however, bioactive steroidal alkaloids, including cyclopamine, veratramine, veratridine, and verazine were identified in its extract. The structural elucidation and bioactivity assessment of steroidal alkaloids in lesser abundance within the extract of V. parviflorum may yield potent Hh pathway inhibitors. This review seeks to consolidate the botanical and phytochemical information regarding V. parviflorum.

HDAC6 inhibition decreases leukemic stem cell expansion driven by Hedgehog hyperactivation by restoring primary ciliogenesis.

Aberrant activation of the Hh pathway promotes cell proliferation and multi-drug resistance (MDR) in several cancers, including Acute Myeloid Leukemia (AML). Notably, only one Hh inhibitor, glasdegib, has been approved for AML treatment, and most patients eventually relapse, highlighting the urgent need to discover new therapeutic targets. Hh signal is transduced through the membrane of the primary cilium, a structure expressed by non-proliferating mammalian cells, whose stabilization depends on the activity of HDAC6. Here we describe a positive correlation between Hh, HDAC6, and MDR genes in a cohort of adult AML patients, human leukemic cell lines, and a zebrafish model of Hh overexpression. The hyper-activation of Hh or HDAC6 in zebrafish drove the increased proliferation of hematopoietic stem and progenitor cells (HSPCs). Interestingly, this phenotype was rescued by inhibition of HDAC6 but not of Hh. Also, in human leukemic cell lines, a reduction in vitality was obtained through HDAC6, but not Hh inhibition. Our data showed the presence of a cross-talk between Hh and HDAC6 mediated by stabilization of the primary cilium, which we detect for the first time in zebrafish HSPCs. Inhibition of HDAC6 activity alone or in combination therapy with the chemotherapeutic agent cytarabine, efficiently rescued the hematopoietic phenotype. Our results open the possibility to introduce HDAC6 as therapeutic target to reduce proliferation of leukemic blasts in AML patients.

Expression of the hedgehog signalling pathway and the effect of inhibition at the level of smoothened in canine osteosarcoma cell lines.

Osteosarcoma (OSA) is the most common malignant bone cancer in dogs. Canine and human OSA share several features, including tumour environments, response to traditional treatment, and several molecular pathways. Hedgehog (Hh) signalling is known to contribute to tumorigenesis and progression of various cancers, including human OSA. This study aimed to identify the role of the Hh signalling pathway in canine OSA cell lines, including Abrams, D17, and Moresco, focusing on the signal transducer Smoothened (SMO). mRNA and protein levels of Hh pathway components, including SHH, IHH, SMO, and PTCH1, were aberrant in all examined OSA cell lines compared with canine osteoblast cells. The SMO inhibitor cyclopamine significantly decreased cell viability and colony-forming ability in the canine OSA cell lines in a dose-dependent manner. Moresco cells, which expressed the highest level of SMO protein, were the most sensitive to the anticancer effect of cyclopamine among the three canine OSA cell lines tested. Hh downstream target gene and protein expression in canine OSA cell lines were downregulated after cyclopamine treatment. In addition, cyclopamine significantly increased apoptotic cell death in Abrams and Moresco cells. The findings that Hh/SMO is activated in canine OSA cell lines and cyclopamine suppresses OSA cell survival via inhibition of SMO suggest that the Hh/SMO signalling pathway might be a novel therapeutic target for canine OSA.

Expression levels of sonic hedgehog pathway genes and their targets are upregulated in early clear cell renal cell carcinoma.

Clear cell renal cell carcinoma (ccRCC) is the most common and aggressive subtype of kidney cancer, with high mortality rates worldwide. The sonic hedgehog (SHH) molecular cascade is altered in various malignancies in tumorigenesis, and several SHH pathway inhibitors have been considered as potential anticancer drugs. The aim of the present study was to determine the expression profile of SHH signaling components and their target genes in ccRCC. Additionally, the present study examined the effects of SHH pathway inhibitory drugs (RU­SKI43, cyclopamine and GLI­antagonist 61) on cell viability, cell cycle progression, expression levels of SHH target genes and migration ability in 786­O, ACHN and HK2 cells. The study also included paired tumor and normal samples from 62 patients with ccRCC. The mRNA levels in clinical samples and cell lines were measured via reverse transcription­quantitative PCR. Cell viability was examined using a sulforhodamine B assay. Flow cytometry was used to investigate cell cycle progression and the migratory rate of cells was assessed using a wound healing assay. High mRNA levels of SHH, smoothened (SMO), glioma­associated zinc finger protein (GLI)1­3, BCL2 apoptosis regulator (BCL2), MYC proto­oncogene (MYC), vascular endothelial growth factor A (VEGFA) and cyclin D1 (CCND1) were observed in the tumor tissues, especially in early ccRCC, according to the TNM stage or World Health Organization/International Society of Urological Pathology (ISUP) grade. High expression levels of VEGFA, as well as low CCND1 mRNA expression, were associated with short overall survival, and increased VEGFA expression was an independent prognostic factor of a poor outcome in patients with advanced ISUP grade (Cox hazard ratio test). Cyclopamine treatment was found to arrest 786­O cells in the G2/M phase and decreased the expression levels of GLI1, BCL2, VEGFA and CCND1. RU­SKI43 inhibited cell migration and decreased the expression levels of BCL2, MYC and CCND1 in ACHN cells. Overall, the results of the present study suggested that SHH signaling may be involved in the early development of ccRCC, and the expression levels of CCND1 and VEGFA may serve as prognostic factors of this disease. Cyclopamine and RU­SKI43 appear to be potential anti­renal cell carcinoma drugs; however, this hypothesis requires verification by further in vivo studies.

The Sonic Hedgehog Pathway Modulates Survival, Proliferation, and Differentiation of Neural Progenitor Cells under Inflammatory Stress In Vitro.

The Sonic Hedgehog protein (Shh) has been extensively researched since its discovery in 1980. Its crucial role in early neurogenesis and endogenous stem cells of mature brains, as well as its recently described neuroprotective features, implicate further important effects on neuronal homeostasis. Here, we investigate its potential role in the survival, proliferation, and differentiation of neural precursors cells (NPCs) under inflammatory stress as a potential adjunct for NPC-transplantation strategies in spinal cord injury (SCI) treatment. To this end, we simulated an inflammatory environment in vitro using lipopolysaccharide (LPS) and induced the Shh-pathway using recombinant Shh or blocked it using Cyclopamine, a potent Smo inhibitor. We found that Shh mediates the proliferation and neuronal differentiation potential of NPCs in vitro, even in an inflammatory stress environment mimicking the subacute phase after SCI. At the same time, our results indicate that a reduction of the Shh-pathway activation by blockage with Cyclopamine is associated with reduced NPC-survival, reduced neuronal differentiation and increased astroglial differentiation. Shh might thus, play a role in endogenous NPC-mediated neuroregeneration or even be a potent conjunct to NPC-based therapies in the inflammatory environment after SCI.

Fourteen-year study of the management of the odontogenic keratocyst. Are adjunctive therapies all they are cut out to be?

Odontogenic keratocysts (OKC) are benign, developmental, locally-aggressive odontogenic cystic lesions with a high risk of recurrence. As such, the most effective treatment modalities remain controversial. The mainstay of treatment remains enucleation with or without decompression. The use of adjunctive therapies is widely reported. Our aim was to review our experience of OKCs and therefore identify the treatment modality, if there is any single one, with the lowest rate of recurrence. We also aimed to identify any common themes linking those patients experiencing cystic recurrence. Data were collected on 50 patients treated at UHCW NHS Trust over a 14-year period (2005-2018) via an anonymised database. Surgical pathways were analysed, including details of the location of the cysts and the use of adjunctive therapies, namely; mechanical debridement, cryotherapy, and the use of Carnoy's solution. Fifty-six keratocysts, both primary (91%, n = 51) and recurrent (9%, n = 5) were included. A total of 6% of patients had a pre-existing diagnosis of Gorlin-Goltz Syndrome (n = 3). Enucleation was performed in an approximately 3:1 ratio to decompression with secondary enucleation (n = 41:15). Twenty-seven percent of patients had adjunctive therapies (n = 15). There was a 12% recurrence rate (n = 6) found only within the group of primary cysts that had been enucleated only. Notably, there were no recurrences in those cysts that had undergone adjunctive therapy. None of the cysts that underwent initial decompression or marsupialisation recurred. Following surgical intervention, no tertiary recurrent cysts were detected postoperatively. This study demonstrated the advantage of establishing a correct diagnosis prior to definitive treatment. Initial decompression in selected patients followed by enucleation, along with adjunctive therapies showed a benefit in reducing recurrences. However, in the absence of high-quality evidence for the most effective management of odontogenic keratocysts, finding a common approach will remain controversial.

[G-protein coupled receptor Smo positively regulates proliferation and migration of adult neural stem cells in vitro].

OBJECTIVE: To investigate the role of G-protein coupled receptor Smoothened (Smo) in regulating proliferation and migration of adult neural stem cells (ANSCs) and explore the underlying mechanism. METHODS: Cultured ANSCs were treated with purmorphamine (PM, an agonist of Smo) or cyclopamine (CPM, an inhibitor of Smo), and the changes in cell proliferation migration abilities were assessed using cell counting kit-8 (CCK8) assay and wound healing assay, respectively. The mRNA expressions of membrane receptor Patched 1 (Ptch1), Smo, glioma-associated oncogene homolog 1 (Gli1), axon guidance cue slit1 (Slit1) and brain-derived neurotrophic factor (BDNF) in the treated cells were detected using real-time quantitative PCR (RT-PCR). RESULTS: PM significantly promoted the proliferation (P < 0.01) and migration of ANSCs (P < 0.01), and up-regulated the mRNA expressions of Ptch1, Smo, Gli1, Slit1 and BDNF. Treatment with CPM significantly inhibited the proliferation and migration of ANSCs. CONCLUSION: Modulating Smo activity can positively regulate the proliferation and migration of ANSCs possibly by regulating the expressions of BDNF and Slit1.

Review: Veratrum californicum Alkaloids.

Veratrum spp. grow throughout the world and are especially prevalent in high mountain meadows of North America. All parts of Veratrum plants have been used for the treatment of ailments including injuries, hypertension, and rheumatic pain since as far back as the 1600s. Of the 17-45 Veratrum spp., Veratrum californicum alkaloids have been proven to possess favorable medicinal properties associated with inhibition of hedgehog (Hh) pathway signaling. Aberrant Hh signaling leads to proliferation of over 20 cancers, including basal cell carcinoma, prostate and colon among others. Six of the most well-studied V. californicum alkaloids are cyclopamine (1), veratramine (2), isorubijervine (3), muldamine (4), cycloposine (5), and veratrosine (6). Recent inspection of the ethanolic extract from V. californicum root and rhizome via liquid chromatography-mass spectrometry has detected up to five additional alkaloids that are proposed to be verazine (7), etioline (8), tetrahydrojervine (9), dihydrojervine (10), 22-keto-26-aminocholesterol (11). For each alkaloid identified or proposed in V. californicum, this review surveys literature precedents for extraction methods, isolation, identification, characterization and bioactivity to guide natural product drug discovery associated with this medicinal plant.