Sources and control of impurity during one-pot enzymatic production of dehydroepiandrosterone.

Dehydroepiandrosterone (DHEA) has a promising market due to its capacity to regulate human hormone levels as well as preventing and treating various diseases. We have established a chemical esterification coupled biocatalytic-based scheme by lipase-catalyzed 4-androstene-3,17-dione (4-AD) hydrolysis to obtain the intermediate product 5-androstene-3,17-dione (5-AD), which was then asymmetrically reduced by a ketoreductase from Sphingomonas wittichii (SwiKR). Co-enzyme required for KR is regenerated by a glucose dehydrogenase (GDH) from Bacillus subtilis. This scheme is more environmentally friendly and more efficient than the current DHEA synthesis pathway. However, a significant amount of 4-AD as by-product was detected during the catalytic process. Focused on the control of by-products, we investigated the source of 4-AD and identified that it is mainly derived from the isomerization activity of SwiKR and GDH. Increasing the proportion of glucose in the catalytic system as well as optimizing the catalytic conditions drastically reduced 4-AD from 24.7 to 6.5% of total substrate amount, and the final yield of DHEA achieved 40.1 g/L. Furthermore, this is the first time that both SwiKR and GDH have been proved to be promiscuous enzymes with dehydrogenase and ketosteroid isomerase (KSI) activities, expanding knowledge of the substrate diversity of the short-chain dehydrogenase family enzymes. KEY POINTS: • A strategy of coupling lipase, ketoreductase, and glucose dehydrogenase in producing DHEA from 4-AD • Both SwiKR and GDH are identified with ketosteroid isomerase activity. • Development of catalytic strategy to control by-product and achieve highly selective DHEA production.

Gate-Controlled Quantum Interference Effects in a Clean Single-Wall Carbon Nanotube p-n Junction.

The precise control and deep understanding of quantum interference in carbon nanotube (CNT) devices are particularly crucial not only for exploring quantum coherent phenomena in clean one-dimensional electronic systems, but also for developing carbon-based nanoelectronics or quantum devices. Here, we construct a double split-gate structure to explore the Aharonov-Bohm (AB) interference effect in individual single-wall CNT p-n junction devices. For the first time, we achieve the AB modulation of conductance with coaxial magnetic fields as low as 3 T, where the flux through the tube is much smaller than the flux quantum. We further demonstrate direct electric-field control of the nonmonotonic magnetoconductance through a gate-tunable built-in electric field, which can be quantitatively understood in combination with the AB phase effect and Landau-Zener tunneling in a CNT p-n junction. Moreover, the nonmonotonic magnetoconductance behavior can be strongly enhanced in the presence of Fabry-Pérot resonances. Our Letter paves the way for exploring and manipulating quantum interference effects with combining magnetic and electric field controls.

Erratum: Estrogen receptor α/prolactin receptor bilateral crosstalk promotes bromocriptine resistance in prolactinomas: Erratum.

[This corrects the article DOI: 10.7150/ijms.51176.].

Structural and Biochemical Analysis Reveals Catalytic Mechanism of Fucoidan Lyase from Flavobacterium sp. SA-0082.

Fucoidans represent a type of polyanionic fucose-containing sulfated polysaccharides (FCSPs) that are cleaved by fucoidan-degrading enzymes, producing low-molecular-weight fucoidans with multiple biological activities suitable for pharmacological use. Most of the reported fucoidan-degrading enzymes are glycoside hydrolases, which have been well studied for their structures and catalytic mechanisms. Little is known, however, about the rarer fucoidan lyases, primarily due to the lack of structural information. FdlA from Flavobacterium sp. SA-0082 is an endo-type fucoidan-degrading enzyme that cleaves the sulfated fuco-glucuronomannan (SFGM) through a lytic mechanism. Here, we report nine crystal structures of the catalytic N-terminal domain of FdlA (FdlA-NTD), in both its wild type (WT) and mutant forms, at resolutions ranging from 1.30 to 2.25 Å. We show that the FdlA-NTD adopts a right-handed parallel ß-helix fold, and possesses a substrate binding site composed of a long groove and a unique alkaline pocket. Our structural, biochemical, and enzymological analyses strongly suggest that FdlA-NTD utilizes catalytic residues different from other ß-helix polysaccharide lyases, potentially representing a novel polysaccharide lyase family.

Toward an Intelligent Synthesis: Monitoring and Intervening in the Catalytic Growth of Carbon Nanotubes.

The bottom-up approach to directly synthesizing low-dimensional materials with outstanding performance has extended the material basis for the next generation integrated circuit industry. All the low-dimensional semiconductors, metals, dielectrics, and their heterojunctions are very promising bricks to build faster and more efficient chips because of their atomically smooth surface and interfaces. The greatest challenge in the synthesis of nanomaterials is how to precisely control the structure, crystalline orientation, defects, dimensions, etc. In past decades, both the methodology and the mechanism of synthesis have been systematically investigated to improve the controllability. However, few studies focused on sensing the synthesis processes in situ and responding to the synthesis immediately. Here, we propose the concept of intelligent synthesis in which the final product can be automatically fine-controlled by a closed loop including in situ monitoring and real-time interventions. As a model system, a high-temperature-tolerant circuit is fabricated on the single-walled carbon nanotube (SWCNT) growth substrate for sensing and responding to the synthesis processes. As a result, either highly pure semiconducting (s-) SWCNT arrays or metallic-semiconducting (m-s) junction arrays with different junction positions is simply synthesized by programming the responding signal. The intelligent synthesis shows much higher efficiency and controllability compared to conventional methods and will lead to the next leap in nanotechnology.

Estrogen receptor α/prolactin receptor bilateral crosstalk promotes bromocriptine resistance in prolactinomas.

Prolactinomas are the most common type of functional pituitary adenoma. Although bromocriptine is the preferred first line treatment for prolactinoma, resistance frequently occurs, posing a prominent clinical challenge. Both the prolactin receptor (PRLR) and estrogen receptor α (ERα) serve critical roles in the development and progression of prolactinomas, and whether this interaction between PRLR and ERα contributes to bromocriptine resistance remains to be clarified. In the present study, increased levels of ERα and PRLR protein expression were detected in bromocriptine-resistant prolactinomas and MMQ cells. Prolactin (PRL) and estradiol (E2) were found to exert synergistic effects on prolactinoma cell proliferation. Furthermore, PRL induced the phosphorylation of ERα via the JAK2-PI3K/Akt-MEK/ERK pathway, while estrogen promoted PRLR upregulation via pERα. ERα inhibition abolished E2-induced PRLR upregulation and PRL-induced ERα phosphorylation, and fulvestrant, an ERα inhibitor, restored pituitary adenoma cell sensitivity to bromocriptine by activating JNK-MEK/ERK-p38 MAPK signaling and cyclin D1 downregulation. Collectively, these data suggest that the interaction between the estrogen/ERα and PRL/PRLR pathways may contribute to bromocriptine resistance, and therefore, that combination treatment with fulvestrant and bromocriptine (as opposed to either drug alone) may exert potent antitumor effects on bromocriptine-resistant prolactinomas.

Overexpression and Biochemical Characterization of an Endo-α-1,4-polygalacturonase from Aspergillus nidulans in Pichia pastoris.

Pectinases have many applications in the industry of food, paper, and textiles, therefore finding novel polygalacturonases is required. Multiple sequence alignment and phylogenetic analysis of AnEPG (an endo-α-1,4-polygalacturonase from Aspergillus nidulans) and other GH 28 endo-polygalacturonases suggested that AnEPG is different from others. AnEPG overexpressed in Pichia pastoris was characterized. AnEPG showed the highest activity at pH 4.0, and exhibited moderate activity over a narrow pH range (pH 2.0-5.0) and superior stability in a wide pH range (pH 2.0-12.0). It displayed the highest activity at 60 °C, and retained >42.2% of maximum activity between 20 and 80 °C. It was stable below 40 °C and lost activity very quickly above 50 °C. Its apparent kinetic parameters against PGA (polygalacturonic acid) were determined, with the Km and kcat values of 8.3 mg/mL and 5640 µmol/min/mg, respectively. Ba2+ and Ni2+ enhanced activity by 12.2% and 9.4%, respectively, while Ca2+, Cu2+, and Mn2+ inhibited activity by 14.8%, 12.8%, and 10.2% separately. Analysis of hydrolysis products by AnEPG proved that AnEPG belongs to an endo-polygalacturonase. Modelled structure of AnEPG by I-TASSER showed structural characteristics of endo-polygalacturonases. This pectinase has great potential to be used in food industry and as feed additives.

SmpB down-regulates proton-motive force for the persister tolerance to aminoglycosides in Aeromonas veronii.

Bacterial persisters comprise a small fraction of phenotypically heterogeneous variants with transient capability for survival when exposed to high concentrations of antibiotic. In aquatic pathogenic bacteria Aeromonas veronii, Small Protein B (SmpB), the core factor of trans-translation system, was identified as a new persistence-related gene. The SmpB deletion exhibited a higher susceptibility and lower persister cell formation under aminoglycosides antibiotics pressure compared with wild type. The transcriptional and translational activities of smpB gene were significantly enhanced by the gentamicin challenge in exponential phase, but not changed in stationary phase. The transcriptomic analysis revealed that the smpB deletion stimulated the production of proton-motive force (PMF). The cell survival induced by carbonyl cyanide m-chlorophenyl hydrazone (CCCP) further verified that SmpB variation affected the quantities of PMF. Taken together, these results uncovered a novel mechanism of persister formation mediated by SmpB under aminoglycosides treatments.

Natural variation in SSW1 coordinates seed growth and nitrogen use efficiency in Arabidopsis.

Seed size is controlled not only by intrinsic genetic factors but also by external environmental signals. Here, we report a major quantitative trait locus (QTL) gene for seed size and weight on chromosome 1 (SSW1) in Arabidopsis, and we found SSW1 acts maternally to positively regulate seed size. Natural variation in SSW1 contains three types of alleles. The SSW1Cvi allele produces larger seeds with more amino acid and storage protein contents than the SSW1Ler allele. SSW1Cvi displays higher capacity for amino acid transport than SSW1Ler due to the differences in transport efficiency. Under low nitrogen supply, the SSW1Cvi allele exhibits increased seed yield and nitrogen use efficiency (NUE). Locations of natural variation alleles of SSW1 are associated with local soil nitrogen contents, suggesting that SSW1 might contribute to geographical adaptation in Arabidopsis. Thus, our findings reveal a mechanism that coordinates seed growth and NUE, suggesting a potential target for improving seed yield and NUE in crops.

Evaluation of coupling coordination development between digital economy and green finance: Evidence from 30 provinces in China.

The convergence of China's digital economy and green finance holds great significance for fostering a sustainable and high-quality developmental path. However, existing studies have not explored the coupling coordination development between these two crucial subsystems. To bridge this gap, this paper employs a modified coupling coordination degree (CCD) model to assess and affirm the coupling coordination degree between the digital economy and green finance across 30 provinces in China from 2015-2021. Based on degree results, provinces are classified into three clusters by using K-means and hierarchical clustering algorithm. Our findings unveil that the current level of coupling coordination development in China is at a primary coordination stage. Although regional disparities significantly exist, the overall level of coordination remains steadily increasing, with the eastern region outperforming the western region. Additionally, we determine that the COVID-19 pandemic's disruption on the coupling coordination development of these systems has been limited. This research sheds light on the evolution of coupling systems and offers practical recommendations for strengthening the coordinated development of the digital economy and green finance.

Radioactivity level evaluation in granite building raw material production area in Cenxi, China.

This study was conducted to evaluate the radioactivity level in the granite building raw material production area in Cenxi, China. Natural radionuclide concentrations, γ absorbed dose rates (ADRs) and radon exhalation rates were measured in the area. The spatial distribution of natural radionuclides, γ ADR and radon exhalation rate were mapped with GPS information. The radioactivity levels in the study area were analysed based on the descriptive statistics and frequency distribution of measurement data. According to the Chinese standard, the granite raw materials used for building and decoration in this region were classified based on their radiological hazards. In addition, radiation protection measures were proposed for the mining of local granite building materials products as well as environmental protection around the work area and resident safety.

Impact of Toll-like Receptor 4 Expression on Inflammatory Responses Related to Premature Membrane Rupture Induced by Lipopolysaccharide.

OBJECTIVE: This study aimed to determine the mechanism through which the expression of Toll-like receptor 4 (TLR4) influences the lipopolysaccharide (LPS)-induced inflammatory response, a condition that is associated with premature rupture of membranes (PROM). METHODS: Human myeloid leukemia mononuclear cells (THP-1) were employed as the experimental model. These cells were treated with LPS and the TLR4 inhibitor CLI-095 and subsequently divided into three groups. A range of assays were utilized, including methyl thiazolyl tetrazole (MTT) assay, real-time fluorescence quantitative polymerase chain reaction (RT-qPCR) for measuring TLR4 and tumor necrosis factor α (TNF-α) mRNA levels, double antibody sandwich enzyme-linked immunosorbent assay (ELISA) for assessing monocyte chemoattractant protein 1 (MCP-1) and matrix metalloproteinase 9 (MMP-9), as well as secretion levels of interleukin (IL)-6 and IL-1ß. And western blotting was used to detect the expression of extracellular signal-regulated kinase (ERK) and nuclear factor κB (NF-κB) p65, which are components of the TLR4 downstream signaling pathway. RESULTS: The LPS-induced proliferation of THP-1 cells was significantly inhibited (p < 0.05) when compared with normal THP-1 cells. Moreover, LPS also promoted TLR4 mRNA and protein expression levels, TNF-α mRNA expression, secretion of inflammatory factors, and phosphorylation of ERK and NF-κB p65 proteins (p < 0.05). On the other hand, administration of the TLR4 inhibitor CLI-095 significantly inhibited the expression of TLR4 mRNA and protein. It also effectively increased the proliferative activity of THP-1 cells and inhibited the secretion of TNF-α and inflammatory factors, as well as the phosphorylation of ERK and NF-κB p65 proteins (p < 0.05). CONCLUSIONS: In summary, suppressing TLR4 expression can mitigate inflammatory responses, thereby reducing the likelihood of premature rupture of membranes during pregnancy, which is often triggered by such inflammation.

A glutamate receptor-like gene is involved in ABA-mediated growth control in Physcomitrium (Physcomitrella) patens.

Plant glutamate receptor homologs (GLRs), which function as key calcium channels, play pivotal roles in various developmental processes as well as stress responses. The moss Physcomitrium patens, a representative of the earliest land plant lineage, possess multiple pathways of hormone signaling for coordinating growth and adaptation responses. However, it is not clear whether GLRs are connected to hormone-mediated growth control in the moss. In this study, we report that one of the two GLRs in P. patens, PpGLR1, involves in abscisic acid (ABA)-mediated growth regulation. ABA represses the growth of wild-type moss, and intriguingly, the PpGLR1 transcript levels are significantly increased in response to ABA treatment, based on both gene expression and the PpGLR1pro::GUS reporter results. Furthermore, the growth of Ppglr1 knockout moss mutants is hypersensitive to ABA treatment. These results suggest that PpGLR1 plays a critical role in ABA-mediated growth regulation, which provide useful information for our further investigation of the regulatory mechanism between Ca2+ signal and ABA in moss growth control.

Cbl upregulates cysH for hydrogen sulfide production in Aeromonas veronii.

Endogenous hydrogen sulfide (H2S) is generated in many metabolism pathways, and has been recognized as a second messenger against antibiotics and reactive oxygen species (ROS). In Aeromonas veronii, Small Protein B (SmpB) plays an important role in resisting stress. The absence of smpB could trigger sulfate assimilation pathway to adapt the nutrient deficiency, of which was mediated by up-regulation of cbl and cys genes and followed with enhancing H2S production. To figure out the mutual regulations of cbl and cys genes, a series of experiments were performed. Compared with the wild type, cysH was down-regulated significantly in cbl deletion by qRT-PCR. The fluorescence analysis further manifested that Cbl had a positive regulatory effect on the promoter of cysJIH. Bacterial one-hybrid analysis and electrophoretic mobility shift assay (EMSA) verified that Cbl bound with the promoter of cysJIH. Collectively, the tolerance to adversity could be maintained by the production of H2S when SmpB was malfunctioned, of which the activity of cysJIH promoter was positively regulated by upstream Cbl protein. The outcomes also suggested the enormous potentials of Aeromonas veronii in environmental adaptability.

Water yield variation with elevation, tree age and density of larch plantation in the Liupan Mountains of the Loess Plateau and its forest management implications.

Large scale afforestation mainly for erosion control or timber production and a very strict logging ban policy in recent decades led to many over-dense stands and remarkable water yield reduction in the dryland region of the Loess Plateau in northwest China. To guide the integrated forest-water management at stand level, a study on the response of water yield from larch (Larix principis-rupprechtii) plantations to key stand structure and site factors was carried out in the Liupan Mountains. Models of leaf area index (LAI) of forest canopy and stand evapotranspiration (ET) in the growing season were developed and fitted. The growing season water yield was calculated based on water budget. The results showed that: (1) The LAI increases with rising tree density firstly quickly and then slowly and finally tending to its maximum; but firstly increases and then decreases with rising tree age and elevation. The LAI model coupling the effects of tree density, age, and elevation works well. (2) The ET model reflecting the coupled effects of precipitation, potential evapotranspiration, LAI and soil moisture can well predict the ET variation. (3) The water yield decreases gradually with rising tree density, but firstly decreases and then increases with rising tree age and elevation. The lowest water yield appears at the age of 30 years and at an elevation of 2420 m. (4) The implications of this study for integrated forest-water management are: defining water yield as the dominant forest service at high or low elevations, but quality timber production as the dominant service at medium elevations; arranging rational thinning for dense forests around the age of 30 years; designing a mosaic distribution of forest ages within watersheds. Applying the study outcomes can promote the integration of water yield management with traditional forest management to ensure the sustainability of water supply in dryland regions.

6 nm super-resolution optical transmission and scattering spectroscopic imaging of carbon nanotubes using a nanometer-scale white light source.

Optical transmission and scattering spectroscopic microscopy at the visible and adjacent wavelengths denote one of the most informative and inclusive characterization methods in material research. Unfortunately, restricted by the diffraction limit of light, it cannot resolve the nanoscale variation in light absorption and scattering, diagnostics of the local inhomogeneity in material structure and properties. Moreover, a large quantity of nanomaterials has anisotropic optical properties that are appealing yet hard to characterize through conventional optical methods. There is an increasing demand to extend the optical hyperspectral imaging into the nanometer length scale. In this work, we report a super-resolution hyperspectral imaging technique that uses a nanoscale white light source generated by superfocusing the light from a tungsten-halogen lamp to simultaneously obtain optical transmission and scattering spectroscopic images. A 6-nm spatial resolution in the visible to near-infrared wavelength regime (415-980 nm) is demonstrated on an individual single-walled carbon nanotube (SW-CNT). Both the longitudinal and transverse optical electronic transitions are measured, and the SW-CNT chiral indices can be identified. The band structure modulation in a SW-CNT through strain engineering is mapped.

[Effects of environment and canopy structure on stem sap flow in a Larix principis-rupprechtii plantation.] / çŽ¯å¢ƒå’Œå† å±‚ç»“æž„å¯¹åŽåŒ—è½å¶æ¾æž—æ ‘å¹²æ¶²æµçš„å½±å“.

Accurately quantifying the impacts of environmental factors and canopy structure on stem sap flow is of great significance for deeply understanding water use strategies of trees in changing environment. The stem sap flow of Larix principis-rupprechtii plantation was observed using thermal diffusion probes from June to September of 2019 in the Xiangshuihe small watershed of Liupan Mountains, with the meteorological conditions, root-zone soil water content and canopy structure being simultaneously recorded. We first analyzed the relationships of sap flow rate (Jc) to potential evapotranspiration (PET), relative extract water (REW) and canopy leaf area index (LAI), and then quantified their relative contribution to Jc. The results showed that the response of Jc to PET, LAI, and REW conformed to binomial, linearly increase and saturated exponential function, respectively. The Jc model coupling multiple factors was established as a continuous multiplication of the response functions of Jc to PET, REW and LAI, which had good simulation precision. PET was the main factor leading to the difference of Jc in different weather conditions. The average contribution rate of PET had obvious difference in sunny (with a contribution rate of 40.3%), cloudy (4.3%), and rainy days (-26.3%). PET and LAI were the leading factors affecting the Jc variation among months. The ranges of the contribution rates of PET and LAI were from -23.1% to 16.8% and from -12.3% to 11.0%, respectively. The Jc model coupling the multi-factor effect developed in this study could be used to predict Jc, and quantify the impacts of each leading factor, which had the potential to be an effective tool to analyze the water use of trees in the changing environment.

Overexpression and biochemical characterization of a truncated endo-α (1 → 3)-fucoidanase from alteromonas sp. SN-1009.

Endo-fucoidanases are important in structural analysis of fucoidans and preparation of fuco-oligosaccharides. However their enzymological properties and analysis of degradation products are scarcely investigated. Truncated endo-α (1 â†’ 3)-fucoidanase Fda1 (tFda1B from Alteromonas sp. was overexpressed and characterized, showing highest activity at pH 7.0, 35 °C, and 1.0 M NaCl. Its Km and kcat were 3.88 ± 0.81 mg/mL and 0.82 ± 0.17 min-1. Fe3+ and Mn2+ enhanced activity by 100% and 19.5% respectively. Co2+ and Cu2+ completely inactivated tFda1B, whereas Ni2+, Mg2+, Zn2+, Pb2+, Ca2+, Ba2+ and Li+ decreased activity by 58.8%, 56.0%, 50.6%, 47.7%, 28.9%, 15.6% and 37.5%, respectively. Catalytic residues were identified through structure and sequence alignment, and confirmed by mutagenesis. Degradation products of Kjellmaniella crassifolia fucoidan by tFda1B were characterized by LC-ESI-MS/MS, confirming tFda1B belongs to endo-(1 â†’ 3)-fucoidanases, and backbone of K. crassifolia fucoidan is 1 â†’ 3 fucoside linkage. This endo-α (1 â†’ 3)-fucoidanase would be useful for elucidating fucoidan structures, and be used as a food enzyme.

GASC1 promotes glioma progression by enhancing NOTCH1 signaling.

Recent studies have reported that gene amplified in squamous cell carcinoma 1 (GASC1) is involved in the progression of several types of cancer. However, whether GASC1 promotes glioma progression remains unknown. Therefore, the present study aimed to investigate the effect of GASC1 exposure on glioma tumorigenesis. The western blot demonstrated that grade III and IV glioma tissues exhibited a higher mRNA and protein expression of GASC1. Moreover, CD133+ U87 or U251 cells from magnetic cell separation exhibited a higher GASC1 expression. Invasion Transwell assay, clonogenic assay and wound healing assay have shown that GASC1 inhibition using a pharmacological inhibitor and specific short hairpin (sh)RNA suppressed the invasive, migratory and tumorsphere forming abilities of primary culture human glioma cells. Furthermore, GASC1­knockdown decreased notch receptor (Notch) responsive protein hes family bHLH transcription factor 1 (Hes1) signaling. GASC1 inhibition reduced notch receptor 1 (NOTCH1) expression, and a NOTCH1 inhibitor enhanced the effects of GASC1 inhibition on the CD133+ U87 or U251 cell tumorsphere forming ability, while NOTCH1 overexpression abrogated these effects. In addition, the GASC1 inhibitor caffeic acid and/or the NOTCH1 inhibitor DAPT (a γ­Secretase Inhibitor), efficiently suppressed the human glioma xenograft tumors. Thus, the present results demonstrated the importance of GASC1 in the progression of glioma and identified that GASC1 promotes glioma progression, at least in part, by enhancing NOTCH signaling, suggesting that GASC1/NOTCH1 signaling may be a potential therapeutic target for glioma treatment.

Pimozide augments bromocriptine lethality in prolactinoma cells and in a xenograft model via the STAT5/cyclin D1 and STAT5/Bcl­xL signaling pathways.

As hyperprolactinemia is observed in patients with bromocriptine­resistant prolactinoma, prolactin (PRL) has been implicated in the development of bromocriptine resistance. Since PRL primarily mediates cell survival and drug resistance via the Janus kinase­2 (JAK2)/signal transducer and activator of transcription 5A (STAT5) signaling pathway, the STAT5 inhibitor, pimozide, may inhibit cell proliferation and reverse bromocriptine resistance in prolactinoma cells. In the present study, compared with bromocriptine or pimozide alone, the combination of pimozide and bromocriptine exerted enhanced reduction in cell growth and proliferation, and increased apoptosis and cell cycle arrest in bromocriptine­resistant prolactinoma cells. A reduction in phospho­STAT5, cyclin D1 and B­cell lymphoma extra­large (Bcl­xL) expression levels were observed in cells treated with the combination of drugs. In addition, pimozide suppressed spheroid formation of human pituitary adenoma stem­like cells, and reduced the protein expression of the cancer stem cell markers, CD133 and nestin. Pimozide did not exert any additional antitumor activity in STAT5­knockdown primary culture cells of human bromocriptine­resistant prolactinomas. Furthermore, Pimozide combined with bromocriptine treatment significantly reduced human prolactinoma xenograft growth. Western blot and immunohistochemical analyses also demonstrated significant inhibition of cell proliferation and stem cell marker proteins in vivo. Collectively, these data indicated that pimozide treatment reduced prolactinoma growth by targeting both proliferating cells and stem cells, at least in part, by inhibiting the STAT5/Bcl­xL and STAT5/cyclin D1 signaling pathways.