Enhanced Golic+: highly effective CRISPR gene targeting and transgene HACKing in Drosophila.

Gene targeting is an incredibly valuable technique. Sometimes, however, it can also be extremely challenging for various intrinsic reasons (e.g. low target accessibility or nature/extent of gene modification). To bypass these barriers, we designed a transgene-based system in Drosophila that increases the number of independent gene targeting events while at the same time enriching for correctly targeted progeny. Unfortunately, with particularly challenging gene targeting experiments, our original design yielded numerous false positives. Here, we deliver a much-improved technique, named Enhanced Golic+ (E-Golic+). E-Golic+ incorporates genetic modifications to tighten lethality-based selection while simultaneously boosting efficiency. With E-Golic+, we easily achieve previously unattainable gene targeting. Additionally, we built an E-Golic+-based, high-efficiency genetic pipeline for transgene swapping. We demonstrate its utility by transforming GAL4 enhancer-trap lines into tissue-specific Cas9-expressing lines. Given the superior efficiency, specificity and scalability, E-Golic+ promises to expedite development of additional sophisticated genetic/genomic tools in Drosophila.

Contribution of AMPA Receptor-Mediated LTD in LA/BLA-CeA Pathway to Comorbid Aversive and Depressive Symptoms in Neuropathic Pain.

Comorbid anxiety and depressive symptoms in chronic pain are a common health problem, but the underlying mechanisms remain unclear. Previously, we have demonstrated that sensitization of the CeA neurons via decreased GABAergic inhibition contributes to anxiety-like behaviors in neuropathic pain rats. In this study, by using male Sprague Dawley rats, we reported that the CeA plays a key role in processing both sensory and negative emotional-affective components of neuropathic pain. Bilateral electrolytic lesions of CeA, but not lateral/basolateral nucleus of the amygdala (LA/BLA), abrogated both pain hypersensitivity and aversive and depressive symptoms of neuropathic rats induced by spinal nerve ligation (SNL). Moreover, SNL rats showed structural and functional neuroplasticity manifested as reduced dendritic spines on the CeA neurons and enhanced LTD at the LA/BLA-CeA synapse. Disruption of GluA2-containing AMPAR trafficking and endocytosis from synapses using synthetic peptides, either pep2-EVKI or Tat-GluA2(3Y), restored the enhanced LTD at the LA/BLA-CeA synapse, and alleviated the mechanical allodynia and comorbid aversive and depressive symptoms in neuropathic rats, indicating that the endocytosis of GluA2-containing AMPARs from synapses is probably involved in the LTD at the LA/BLA-CeA synapse and the comorbid aversive and depressive symptoms in neuropathic pain in SNL-operated rats. These data provide a novel mechanism for elucidating comorbid aversive and depressive symptoms in neuropathic pain and highlight that structural and functional neuroplasticity in the amygdala may be important as a promising therapeutic target for comorbid negative emotional-affective disorders in chronic pain.SIGNIFICANCE STATEMENT Several studies have demonstrated the high comorbidity of negative affective disorders in patients with chronic pain. Understanding the affective aspects related to chronic pain may facilitate the development of novel therapies for more effective management. Here, we unravel that the CeA plays a key role in processing both sensory and negative emotional-affective components of neuropathic pain, and LTD at the amygdaloid LA/BLA-CeA synapse mediated by GluA2-containing AMPAR endocytosis underlies the comorbid aversive and depressive symptoms in neuropathic pain. This study provides a novel mechanism for elucidating comorbid aversive and depressive symptoms in neuropathic pain and highlights that structural and functional neuroplasticity in the amygdala may be important as a promising therapeutic target for comorbid negative emotional-affective disorders in chronic pain.

Calycosin inhibited autophagy and oxidative stress in chronic kidney disease skeletal muscle atrophy by regulating AMPK/SKP2/CARM1 signalling pathway.

Skeletal muscle atrophy is a common and serious complication of chronic kidney disease (CKD). Oxidative stress and autophagy are the primary molecular mechanisms involved in muscle atrophy. Calycosin, a major component of Radix astragali, exerts anti-inflammatory, anti-oxidative stress and anti-autophagy effects. We investigated the effects and mechanisms of calycosin on skeletal muscle atrophy in vivo and in vitro. 5/6 nephrectomy (5/6 Nx) rats were used as a model of CKD. We evaluated bodyweight and levels of serum creatinine (SCr), blood urea nitrogen (BUN) and serum albumin (Alb). H&E staining, cell apoptosis, oxidative stress biomarkers, autophagosome and LC3A/B levels were performed and evaluated in skeletal muscle of CKD rat. Calycosin treatment improved bodyweight and renal function, alleviated muscle atrophy (decreased the levels of MuRF1 and MAFbx), increased superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px) activity and reduced malondialdehyde (MDA) levels in skeletal muscle of CKD rats. Importantly, calycosin reduced autophagosome formation, down-regulated the expression of LC3A/B and ATG7 through inhibition of AMPK and FOXO3a, and increased SKP2, which resulted in decreased expression of CARM1, H3R17me2a. Similar results were observed in C2C12 cells treated with TNF-α and calycosin. Our findings showed that calycosin inhibited oxidative stress and autophagy in CKD induced skeletal muscle atrophy and in TNF-α-induced C2C12 myotube atrophy, partially by regulating the AMPK/SKP2/CARM1 signalling pathway.

Transcriptomes of lineage-specific Drosophila neuroblasts profiled by genetic targeting and robotic sorting.

A brain consists of numerous distinct neurons arising from a limited number of progenitors, called neuroblasts in Drosophila. Each neuroblast produces a specific neuronal lineage. To unravel the transcriptional networks that underlie the development of distinct neuroblast lineages, we marked and isolated lineage-specific neuroblasts for RNA sequencing. We labeled particular neuroblasts throughout neurogenesis by activating a conditional neuroblast driver in specific lineages using various intersection strategies. The targeted neuroblasts were efficiently recovered using a custom-built device for robotic single-cell picking. Transcriptome analysis of mushroom body, antennal lobe and type II neuroblasts compared with non-selective neuroblasts, neurons and glia revealed a rich repertoire of transcription factors expressed among neuroblasts in diverse patterns. Besides transcription factors that are likely to be pan-neuroblast, many transcription factors exist that are selectively enriched or repressed in certain neuroblasts. The unique combinations of transcription factors present in different neuroblasts may govern the diverse lineage-specific neuron fates.

Contribution of microglial reaction to increased nociceptive responses in high-fat-diet (HFD)-induced obesity in male mice.

The progressive increase in the prevalence of obesity in the population can result in increased healthcare costs and demands. Recent studies have revealed a positive correlation between pain and obesity, although the underlying mechanisms still remain unknown. Here, we aimed to clarify the role of microglia in altered pain behaviors induced by high-fat diet (HFD) in male mice. We found that C57BL/6CR mice on HFD exhibited enhanced spinal microglial reaction (increased cell number and up-regulated expression of p-p38 and CD16/32), increased tumor necrosis factor-α (TNF-α) mRNA and brain-derived neurotrophic factor (BDNF) protein expression as well as a polarization of spinal microglial toward a pro-inflammatory phenotype. Moreover, we found that using PLX3397 (a selective colony-stimulating factor-1 receptor (CSF1R) kinase inhibitor) to eliminate microglia in HFD-induced obesity mice, inflammation in the spinal cord was rescued, as was abnormal pain hypersensitivity. Intrathecal injection of Mac-1-saporin (a saporin-conjugated anti-mac1 antibody) resulted in a decreased number of microglia and attenuated both mechanical allodynia and thermal hyperalgesia in HFD-fed mice. These results indicate that the pro-inflammatory functions of spinal microglia have a special relevance to abnormal pain hypersensitivity in HFD-induced obesity mice. In conclusion, our data suggest that HFD induces a classical reaction of microglia, characterized by an enhanced phosphorylation of p-38 and increased CD16/32 expression, which may in part contribute to increased nociceptive responses in HFD-induced obesity mice.

SKA3 promotes cell proliferation and migration in cervical cancer by activating the PI3K/Akt signaling pathway.

BACKGROUND: Cervical cancer (CC) is one of the most common cancers among females worldwide. Spindle and kinetochore-associated complex subunit 3 (SKA3), located on chromosome 13q, was identified as a novel gene involved in promoting malignant transformation in cancers. However, the function and underlying mechanisms of SKA3 in CC remain unknown. Using the Oncomine database, we found that expression of SKA3 mRNA is higher in CC tissues than in normal tissues and is linked with poor prognosis. METHODS: In our study, immunohistochemistry showed increased expression of SKA3 in CC tissues. The effect of SKA3 on cell proliferation and migration was evaluated by CCK8, clone formation, Transwell and wound-healing assays in HeLa and SiHa cells with stable SKA3 overexpression and knockdown. In addition, we established a xenograft tumor model in vivo. RESULTS: SKA3 overexpression promoted cell proliferation and migration and accelerated tumor growth. We further identified that SKA3 is involved in regulating cell cycle progression and the PI3K/Akt signaling pathway via RNA-sequencing (RNA-Seq) and gene set enrichment analyses. Western blotting results revealed that SKA3 overexpression increased levels of p-Akt, cyclin E2, CDK2, cyclin D1, CDK4, E2F1 and p-Rb in HeLa cells. Additionally, the use of an Akt inhibitor (GSK690693) significantly reversed the cell proliferation capacity induced by SKA3 overexpression in HeLa cells. CONCLUSIONS: We suggest that SKA3 overexpression contributes to CC cell growth and migration by promoting cell cycle progression and activating the PI3K-Akt signaling pathway, which may provide potential novel therapeutic targets for CC treatment.

Electroacupuncture Treatment Alleviates the Remifentanil-Induced Hyperalgesia by Regulating the Activities of the Ventral Posterior Lateral Nucleus of the Thalamus Neurons in Rats.

Mechanisms underlying remifentanil- (RF-) induced hyperalgesia, a phenomenon that is generally named as opioid-induced hyperalgesia (OIH), still remain elusive. The ventral posterior lateral nucleus (VPL) of the thalamus, a key relay station for the transmission of nociceptive information to the cerebral cortex, is activated by RF infusion. Electroacupuncture (EA) is an effective method for the treatment of pain. This study aimed to explore the role of VPL in the development of OIH and the effect of EA treatment on OIH in rats. RF was administered to rats via the tail vein for OIH induction. Paw withdrawal threshold (PWT) in response to mechanical stimuli and paw withdrawal latency (PWL) to thermal stimulation were tested in rats for the assessment of mechanical allodynia and thermal hyperalgesia, respectively. Spontaneous neuronal activity and local field potential (LFP) in VPL were recorded in freely moving rats using the in vivo multichannel recording technique. EA at 2 Hz frequency (pulse width 0.6 ms, 1-3 mA) was applied to the bilateral acupoints "Zusanli" (ST.36) and "Sanyinjiao" (SP.6) in rats. The results showed that both the PWT and PWL were significantly decreased after RF infusion to rats. Meanwhile, both the spontaneous neuronal firing rate and the theta band oscillation in VPL LFP were increased on day 3 post-RF infusion, indicating that the VPL may promote the development of RF-induced hyperalgesia by regulating the pain-related cortical activity. Moreover, 2 Hz-EA reversed the RF-induced decrease both in PWT and PWL of rats and also abrogated the RF-induced augmentation of the spontaneous neuronal activity and the power spectral density (PSD) of the theta band oscillation in VPL LFP. These results suggested that 2 Hz-EA attenuates the remifentanil-induced hyperalgesia via reducing the excitability of VPL neurons and the low-frequency (theta band) oscillation in VPL LFP.

An Oxalyl-CoA Synthetase Is Involved in Oxalate Degradation and Aluminum Tolerance.

Acyl Activating Enzyme3 (AAE3) was identified to be involved in the catabolism of oxalate, which is critical for seed development and defense against fungal pathogens. However, the role of AAE3 protein in abiotic stress responses is unknown. Here, we investigated the role of rice bean (Vigna umbellata) VuAAE3 in Al tolerance. Recombinant VuAAE3 protein has specific activity against oxalate, with Km = 121 ± 8.2 µm and Vmax of 7.7 ± 0.88 µmol min-1 mg-1 protein, indicating it functions as an oxalyl-CoA synthetase. VuAAE3-GFP localization suggested that this enzyme is a soluble protein with no specific subcellular localization. Quantitative reverse transcription-PCR and VuAAE3 promoter-GUS reporter analysis showed that the expression induction of VuAAE3 is mainly confined to rice bean root tips. Accumulation of oxalate was induced rapidly by Al stress in rice bean root tips, and exogenous application of oxalate resulted in the inhibition of root elongation and VuAAE3 expression induction, suggesting that oxalate accumulation is involved in Al-induced root growth inhibition. Furthermore, overexpression of VuAAE3 in tobacco (Nicotiana tabacum) resulted in the increase of Al tolerance, which was associated with the decrease of oxalate accumulation. In addition, NtMATE and NtALS3 expression showed no difference between transgenic lines and wild-type plants. Taken together, our results suggest that VuAAE3-dependent turnover of oxalate plays a critical role in Al tolerance mechanisms.

BDNF Contributes to Spinal Long-Term Potentiation and Mechanical Hypersensitivity Via Fyn-Mediated Phosphorylation of NMDA Receptor GluN2B Subunit at Tyrosine 1472 in Rats Following Spinal Nerve Ligation.

Previously we have demonstrated that brain-derived neurotrophic factor (BDNF) contributes to spinal long-term potentiation (LTP) and pain hypersensitivity through activation of GluN2B-containing N-methyl-D-aspartate (GluN2B-NMDA) receptors in rats following spinal nerve ligation (SNL). However, the molecular mechanisms by which BDNF impacts upon GluN2B-NMDA receptors and spinal LTP still remain unclear. In this study, we first documented that Fyn kinase-mediated phosphorylation of GluN2B subunit at tyrosine 1472 (pGluN2BY1472) was involved in BDNF-induced spinal LTP and pain hypersensitivity in intact rats. Second, we revealed a co-localization of Fyn and GluN2B-NMDA receptor in cultured dorsal horn neurons, implying that Fyn is a possible intermediate kinase linking BDNF/TrkB signaling with GluN2B-NMDA receptors in the spinal dorsal horn. Furthermore, we discovered that both SNL surgery and intrathecal active Fyn could induce an increased expression of dorsal horn pGluN2BY1472, as well as pain hypersensitivity in response to von Frey filaments stimuli; and more importantly, all these actions were effectively abrogated by pre-treatment with either PP2 or ifenprodil to respectively inhibit Fyn kinase and GluN2B-NMDA receptors activity. Moreover, we found that intrathecal administration of BDNF scavenger TrkB-Fc prior to SNL surgery, could prevent the nerve injury-induced increase of both pFynY420 and pGluN2BY1472 expression, and also inhibit the mechanical allodynia in neuropathic rats. Collectively, these results suggest that Fyn kinase-mediated pGluN2BY1472 is critical for BDNF-induced spinal LTP and pain hypersensitivity in SNL rats. Therefore, the BDNF-Fyn-GluN2B signaling cascade in the spinal dorsal horn may constitute a key mechanism underlying central sensitization and neuropathic pain development after peripheral nerve injury.

Pattern Recognition to Prognosticate Endometrial Cancer: The Science Behind the Art of Office Hysteroscopy-A Retrospective Study.

OBJECTIVE: This study aimed to evaluate a specific glomerular pattern for prognostication of endometrial cancer (EC). MATERIALS AND METHODS: The office hysteroscopy's picture and video of 4197 women were reviewed, 48 women who were suspected of type I EC were analyzed: 26 have glomerular pattern (group 1) and 22 without it (group 2). RESULTS: The histopathological grading after hysterectomy with glomerular pattern had grade 2 or grade 3 disease on final histology (n = 25; 96%). The sensitivity and specificity of this test were 84.6% and 81.8%, respectively, with a likelihood ratio of 4:6 in predicting and prognosticating those women who have high-grade tumor or invasive disease. CONCLUSIONS: This hysteroscopic picture might be used as a novel marker for risk stratification of EC.

Evaluation of ecotourism suitability based on AHP-GIS: Taking Xiaoxiangling area of the Giant Panda National Park and the surrounding communities as an example.

The primary goal of national parks is to protect ecological environment, but also with the functions of scientific research, education, and recreation. Aiming for the realization of universal sharing, we used the analytic hierarchy process (AHP) to construct an ecotourism suitability evaluation system by selecting four factors, including landscape resources, ecological environment carrying capacity, recreation utilization capacity and social condition, taking Xiaoxiangling area of Giant Panda National Park and the surrounding communities as an example. We evaluated the ecotourism suitability based on GIS, and conducted a questionnaire survey of tourists, to propose suggestions on the functional zoning in terms of ecotourism suitability and subjective choice preferences of tourists. The results showed that the ecotourism suitability of the evaluation area could be classified into five levels. The most suitable areas were located nearby the natural landscape resources and far away from the core conservation area, and the least suitable areas distributed at the edge of the core conservation area. According to the results of suitability analysis, the evaluation area was divided into suitable development area, moderate development area, and restricted development area. Combined with the tourist preferences, we divided the recreational activities in the evaluation area into seven activities, namely, ecotourism, eco-camping, science education, leisure vacation, agricultural and animal husbandry culture experience, eco-education, and mountain adventure. These findings could help provide suitable services for different tourists and offer reference for the ecotourism developmental planning of the Xiaoxiangling area of the Giant Panda National Park.

N-Hydroxycinnamide derivatives of osthole presenting genotoxicity and cytotoxicity against human colon adenocarcinoma cells in vitro and in vivo.

Osthole is extracted from the Chinese herbs Cnidium monnieri and Angelica pubescens, and it was found to have antitumor activity in vitro and in vivo. A series of osthole derivatives have been synthesized, and the N-hydroxycinnamide derivatives of osthole, WJ1376-1 and WJ1398-1 were found to have the greatest potential against human colon adenocarcinoma cells. In contrast to the parental osthole, both WJ1376-1 and WJ1398-1 were found to induce multinucleation and polyploidy by microscopic observation and flow cytometry. WJ1376-1 and WJ1398-1 significantly activated ataxia telangiectasia and rad3 related (ATR) kinase, which triggered activation of the checkpoint kinase 2 (Chk2) signaling pathway and then down regulated Cdc25 phosphatase and Cdc2/cyclin B kinase activities. WJ1376-1 and WJ1398-1 also inhibited the phosphorylation of Aurora A kinase, which is associated with important processes during mitosis. The presence of a "comet" DNA fragment and phosphorylation of p53 at Ser 15 clearly indicated that DNA damage occurred with WJ1376-1 and WJ1398-1 treatment. WJ1376-1 and WJ1398-1 ultimately induced apoptosis as evidenced by the upregulation of Bad and activation of caspases-3, -7, and -9. Furthermore, WJ1376-1 and WJ1398-1 also showed a great effect in attenuating tumor growth without affecting the body weight of xenograft nude mice. Taken together, these results suggest that the toxic activities of WJ1376-1 and WJ1398-1 were dissimilar to that of the parental osthole, which can induce cell polyploidy and G2/M cell cycle arrest in colon adenocarcinoma cells and may provide a potential therapeutic target for colon cancer treatment in the future.

Hormone-controlled changes in the differentiation state of post-mitotic neurons.

While we think of neurons as having a fixed identity, many show spectacular plasticity.1-10 Metamorphosis drives massive changes in the fly brain;11,12 neurons that persist into adulthood often change in response to the steroid hormone ecdysone.13,14 Besides driving remodeling,11-14 ecdysone signaling can also alter the differentiation status of neurons.7,15 The three sequentially born subtypes of mushroom body (MB) Kenyon cells (γ, followed by α'/ß', and finally α/ß)16 serve as a model of temporal fating.17-21 γ neurons are also used as a model of remodeling during metamorphosis. As γ neurons are the only functional Kenyon cells in the larval brain, they serve the function of all three adult subtypes. Correspondingly, larval γ neurons have a similar morphology to α'/ß' and α/ß neurons-their axons project dorsally and medially. During metamorphosis, γ neurons remodel to form a single medial projection. Both temporal fate changes and defects in remodeling therefore alter γ-neuron morphology in similar ways. Mamo, a broad-complex, tramtrack, and bric-à-brac/poxvirus and zinc finger (BTB/POZ) transcription factor critical for temporal specification of α'/ß' neurons,18,19 was recently described as essential for γ remodeling.22 In a previous study, we noticed a change in the number of adult Kenyon cells expressing γ-specific markers when mamo was manipulated.18 These data implied a role for Mamo in γ-neuron fate specification, yet mamo is not expressed in γ neurons until pupariation,18,22 well past γ specification. This indicates that mamo has a later role in ensuring that γ neurons express the correct Kenyon cell subtype-specific genes in the adult brain.

Formulation and stability of an extemporaneously compounded propafenone hydrochloride oral suspension for children with arrhythmia.

OBJECTIVES: In children, supraventricular tachycardia is the most common form of arrhythmia, and propafenone is an effective class Ic antiarrhythmic agent used in this population. No suitable paediatric-specific, dosing-flexible preparation is available in Taiwan. The objective of this study was to develop a formulation of propafenone oral suspension prepared from commercially available propafenone tablets and commercially available oral syrup vehicles for related patients. METHODS: An oral suspension of propafenone hydrochloride at a concentration of 10 mg/mL was prepared by mixing finely grounded propafenone hydrochloride tablets and a 1:1 mixture of Ora-Plus and Ora-Sweet. The beyond-use date was determined by analysing the samples stored at room temperature or 2-8℃ at time 0 and on days 7, 14, 21, 28, 35, 42, 56, and 90. Parameters to be inspected included appearance, pH measurement, high-performance liquid chromatography analysis, and microbial limit tests. RESULTS: On the basis of the physicochemical and microbial stability results, the 10 mg/mL oral suspension of propafenone hydrochloride was stable at 2-8℃ and room temperature for at least 90 days. The suspension did not exhibit significant changes in drug concentration or pH level at any time point. Moreover, no apparent changes or microbial contaminations were observed for at least 90 days. CONCLUSIONS: Propafenone hydrochloride in a 10 mg/mL oral suspension prepared by diluting fine powder with a 1:1 mixture of Ora-Plus and Ora-Sweet and stored in high-density polyethylene bottles and has a beyond-use date of 90 days when stored at 2-8℃ or room temperature. This finding enables us to improve the accuracy of dosage administration and reduce the risk of medication errors affecting the paediatric population.

Correlation between gut microbiota and glucagon-like peptide-1 in patients with gestational diabetes mellitus.

BACKGROUND: Gestational diabetes mellitus (GDM) places both the mother and offspring at high risk of complications. Increasing evidence suggests that the gut microbiota plays a role in the pathogenesis of GDM. However, it is still unclear whether the gut microbiota is related to blood biochemical traits, particularly glucagon-like peptide-1 (GLP-1), in GDM patients. AIM: To explore the correlation between the gut microbiota and blood biochemical traits, particularly GLP-1, in GDM patients. METHODS: The V4 region of the 16S ribosomal ribonucleic acid (rRNA) gene was sequenced based on the fecal samples of 35 pregnant women with GDM and was compared to that of 25 pregnant women with normal glucose tolerance (NGT). RESULTS: The results showed that Ruminococcaceae_UCG-002, Ruminococcaceae_UCG-005, Clostri-dium_sensu_stricto_1, and Streptococcus were more abundant in the NGT group than in the GDM group. Bacteroides and Lachnoclostridium were more abundant in the GDM group than in the NGT group. Spearman's correlation analysis was performed to identify the relationships between microbiota genera and blood biochemical traits. Paraprevotella, Roseburia, Faecalibacterium, and Ruminococcaceae_UCG-002 were significantly negatively correlated with glucose. Ruminococcaceae_UCG-002 was significantly negatively correlated with hemoglobin A1c. Bacteroides was significantly positively correlated with glucose. Sutterella, Oscillibacter, and Bifidobacterium were significantly positively correlated with GLP-1. A random forest model showed that 20 specific genera plus glucose provided the best discriminatory power, as indicated by the area under the receiver operating characteristic curve (0.94). CONCLUSION: The results of this study reveal novel relationships between the gut microbiome, blood bio-chemical traits, particularly GLP-1, and GDM status. These findings suggest that some genera are crucial for controlling blood glucose-related indices and may be beneficial for GDM treatment. Alteration in the microbial composition of the gut may potentially serve as a marker for identifying individuals at risk of GDM.

An Unusual Case of Scalp Metastasis from Breast Cancer.

BACKGROUND: The most common sites of breast cancer metastases are the bone, lung, liver, and brain. Scalp involvement in breast cancer metastasis is extraordinarily rare. CASE DESCRIPTION: This study reports a 52-year-old woman who had a history of malignant right breast cancer and underwent a mastectomy. Positron emission tomography/computed tomography revealed a soft tissue nodule measuring 1 × 0.7 cm located subcutaneously on the top left side of the scalp. A scalp mass excision operation was performed with an extended "S"-shaped incision, and the mass was sent for pathology. Immunohistochemistry showed the following results: CK7: +; ER: 2+, 90%; GATA3: +; GCDFP-15: scattered cells+; mammaglobin: -, napsin A: -; and TTF-1: -. These results were consistent with the characteristics of primary right breast cancer, supporting scalp metastasis from breast cancer. CONCLUSIONS: Scalp metastasis from breast cancer is an exceedingly infrequent phenomenon. Close attention should be paid to soft tissue masses in patients with a healthy appearance and in those with a history of malignant cancer. When neurosurgeons operate on the mass, the circumscription and depth of the tumor must be given further attention.

Responses of Primary Afferent Fibers to Acupuncture-Like Peripheral Stimulation at Different Frequencies: Characterization by Single-Unit Recording in Rats.

The pain-relieving effect of acupuncture is known to involve primary afferent nerves (PANs) via their roles in signal transmission to the CNS. Using single-unit recording in rats, we characterized the generation and transmission of electrical signals in Aß and Aδ fibers induced by acupuncture-like stimuli. Acupuncture-like signals were elicited in PANs using three techniques: manual acupuncture (MAc), emulated acupuncture (EAc), and electro-acupuncture (EA)-like peripheral electrical stimulation (PES). The discharges evoked by MAc and EAc were mostly in a burst pattern with average intra-burst and inter-burst firing rates of 90 Hz and 2 Hz, respectively. The frequency of discharges in PANs was correlated with the frequency of PES. The highest discharge frequency was 246 Hz in Aß fibers and 180 Hz in Aδ fibers. Therefore, EA in a dense-disperse mode (at alternating frequency between 2 Hz and 15 Hz or between 2 Hz and 100 Hz) best mimics MAc. Frequencies of EA output >250 Hz appear to be obsolete for pain relief.

Use of sodium glycerophosphate in neonatal parenteral nutrition solutions to increase calcium and phosphate compatibility for preterm infants.

BACKGROUND: Preterm infants require higher calcium and phosphate intake than term infants to facilitate adequate bone growth, but this is rarely met in parenteral nutrition (PN) solution because of the limited solubility of calcium and phosphate. This study aimed to evaluate the solubility of organic phosphate with calcium gluconate in neonatal PN solutions, simulating its clinical use. METHODS: PN solutions were composed of calcium gluconate at 50 mEq/L and sodium glycerophosphate (NaGP) at 25 mmol/L. Another component included 1% or 4% amino acid and 10% or 20% dextrose. For comparison, PN solution composed of potassium phosphate was also evaluated. Each solution was evaluated using the following methods: visual inspection, light obscuration particle count test, and pH measurement. To simulate the clinical condition, the solution was tested after compounding, after being stored at 25 °C for 24 h, and after being stored at 2°C-8°C for 2 or 9 days and subsequently at 25 °C for 24 h. RESULTS: There was no visual deposition in PN solution using NaGP in any of the concentrations and under any stored condition. The solution fulfilled the criteria of physical compatibility as < 25 particles/mL measuring ≥10 µm in diameter and <3 particles/mL measuring ≥25 µm in diameter. On the contrary, visual deposition was evidently noted in PN solution using potassium phosphate after its formulation, and the particle count significantly exceeded the range of physical compatibility. CONCLUSION: NaGP and calcium gluconate have significantly good compatibility in PN solution. The use of NaGP in neonatal PN prevents calcium and phosphorus precipitation, hence increasing their supply to preterm infants in meeting their growth requirement.

Salidroside ameliorates endothelial inflammation and oxidative stress by regulating the AMPK/NF-κB/NLRP3 signaling pathway in AGEs-induced HUVECs.

Endothelial dysfunction plays important roles in vascular dysfunction under diabetic conditions. The generation of advanced glycation end products (AGEs), which can induce inflammation and oxidative stress, is pivotal in endothelial dysfunction. Salidroside, a major active compound in Rhodiola rosea, exerts protective effects against vascular diseases. To study the effects and mechanism of salidroside in diabetes-induced vascular endothelial dysfunction, an in vitro model was established with AGEs-induced human umbilical vein endothelial cells (HUVECs). Then, cell viability, cell apoptosis, pro-inflammatory cytokines and oxidative biomarkers were tested to determine the effects of salidroside at 10, 50 and 100 µM doses on AGEs induced HUVECs. Additionally, RNA-Seq and bioinformatics analyses were used to search for the underlying mechanism of salidroside. The results showed that salidroside promoted cell viability and significantly alleviated cell apoptosis in AGEs-induced HUVECs. Furthermore, salidroside remarkably decreased the levels of the pro-inflammatory cytokines TNF-α, IL-1ß and IL-6 and impeded the expression of VCAM-1 and ICAM-1 induced by AGEs. Additionally, salidroside promoted superoxide dismutase (SOD) activity and increased catalase (CAT) and glutathione peroxidase (GSH-Px) levels while inhibiting the intracellular generation of reactive oxygen species (ROS) and malondialdehyde (MDA) in AGEs-induced HUVECs. Importantly, salidroside alleviated endothelial inflammation and oxidative stress by activating AMPK phosphorylation and inhibiting NF-ĸB p65 and NLRP3 inflammasome activation. Therefore, we used compound C, an accepted AMPK inhibitor, to further demonstrate the mechanism. Interestingly, the phenomenon produced by salidroside was abolished. Our findings suggest that salidroside ameliorates AGEs-induced endothelial inflammation and oxidative stress, partially via the AMPK/NF-κB/NLRP3 signaling pathway.

Function of Drosophila mob2 in photoreceptor morphogenesis.

The Drosophila photoreceptor is a highly polarized cell; a mature photoreceptor cell in Drosophila contains a photosensitive structure (the rhabdomere) and a supporting membrane (stalk) at its apical membrane. In a screen to isolate genes involved in determining stalk and rhabdomere formation, this study has identified the Drosophila mob2 (Dmob2) gene. Dmob2 belongs to a Mob1/phocein domain protein family whose functions are involved in polarized cell growth and asymmetric cell fate determination in yeast. To study the role of Dmob2 in photoreceptor development, we have raised an antibody against the Dmob2 protein. An immunocytochemical study has shown that Dmob2 is mainly localized in the apical membrane of photoreceptor cells during early development. As development proceeds, Dmob2 is gradually confined to the rhabdomere base of the photoreceptor cells. RNA interference (RNAi) for knockdown Dmob2 expression during eye development impairs rhabdomere formation. Our study further shows that the subcellular localization of phosphorylated Moesin and Crumbs in the developing photoreceptor cell is disrupted in Dmob2 RNAi flies. This work thus reports a novel function of Dmob2 in photoreceptor cell development.