Spotlight on porphyrins: Classifications, mechanisms and medical applications.

Photodynamic therapy (PDT) and sonodynamic therapy (SDT) are non-invasive treatment methods with obvious inhibitory effect on tumors and have few side effects, which have been widely concerned and explored by researchers. Sensitizer is the main factor in determining the therapeutic effect of PDT and SDT. Porphyrins, a group of organic compounds widespread in nature, can be activated by light or ultrasound and produce reactive oxygen species. Therefore, porphyrins as sensitizers in PDT have been widely explored and investigated for many years. Herein, we summarize the classical porphyrin compounds and their applications and mechanisms in PDT and SDT. The application of porphyrin in clinical diagnosis and imaging is also discussed. In conclusion, porphyrins have good application prospects in disease treatment as an important part of PDT or SDT, and in clinical diagnosis and imaging.

Menin-MLL1 Interaction Small Molecule Inhibitors: A Potential Therapeutic Strategy for Leukemia and Cancers.

Encoded by the MEN1 gene, menin protein is a fusion protein that is essential for the oncogenic transformation of mixed-lineage leukemia (MLL) and leads to acute leukemia (AL). Therefore, accumulating evidence has demonstrated that inhibition of the high-affinity relationship between menin and mixed-lineage leukemia 1 (MLL1 and KMT2A) is an effective treatment for MLL-rearranged (MLL-r) leukemia in vitro and in vivo. Meanwhile, recent studies found that menin-MLL1 interaction inhibitors exhibited a firm tumor suppressive ability in specific cancer cells, such as prostate cancer, breast cancer, liver cancer, and lung cancer. Overall, it seems to serve as a novel therapeutic means for cancers. Herein, we review the recent progress in exploring the inhibitors of small molecule menin-MLL1 interactions. The molecular mechanisms of these inhibitors' functions and their application prospects in the treatment of AL and cancers are explored.

MEN1 silencing triggers the dysregulation of mTORC1 and MYC pathways in ER+ breast cancer cells.

Menin, encoded by the MEN1 gene, has been identified as a critical factor regulating ESR1 transcription, playing an oncogenic role in ER+ breast cancer (BC) cells. Here, we further dissected the consequences of menin inactivation in ER+ BC cells by focusing on factors within two major pathways involved in BC, mTOR and MYC. MEN1 silencing in MCF7 and T-47D resulted in an increase in phosphor-p70S6K1, phosphor-p85S6K1 and phosphor-4EBP1 expression. The use of an AKT inhibitor inhibited the activation of S6K1 and S6RP triggered by MEN1 knockdown (KD). Moreover, MEN1 silencing in ER+ BC cells led to increased formation of the eIF4E and 4G complex. Clinical studies showed that patients with menin-low breast cancer receiving tamoxifen plus everolimus displayed a trend toward better overall survival. Importantly, MEN1 KD in MCF7 and T-47D cells led to reduced MYC expression. ChIP analysis demonstrated that menin bound not only to the MYC promoter but also to its 5' enhancer. Furthermore, E2-treated MEN1 KD MCF7 cells displayed a decrease in MYC activation, suggesting its role in estrogen-mediated MYC transcription. Finally, expression data mining in tumors revealed a correlation between the expression of MEN1 mRNA and that of several mTORC1 components and targets and a significant inverse correlation between MEN1 and two MYC inhibitory factors, MYCBP2 and MYCT1, in ER+ BC. The current work thus highlights altered mTORC1 and MYC pathways after menin inactivation in ER+ BC cells, providing insight into the crosstalk between menin, mTORC1 and MYC in ER+ BC.

Reduced menin expression leads to decreased ERα expression and is correlated with the occurrence of human luminal B-like and ER-negative breast cancer subtypes.

PURPOSE: Menin, encoded by the MEN1 gene, was recently reported to be involved in breast cancers, though the underlying mechanisms remain elusive. In the current study, we sought to further determine its role in mammary cells. METHODS: Menin expression in mammary lesions from mammary-specific Men1 mutant mice was detected using immunofluorescence staining. RT-qPCR and western blot were performed to determine the role of menin in ERα expression in human breast cancer cell lines. ChIP-qPCR and reporter gene assays were carried out to dissect the action of menin on the proximal ESR1 promoter. Menin expression in female patients with breast cancer was analyzed and its correlation with breast cancer subtypes was investigated. RESULTS: Immunofluorescence staining revealed that early mammary neoplasia in Men1 mutant mice displayed weak ERα expression. Furthermore, MEN1 silencing led to both reduced ESR1 mRNA and ERα protein expression in MCF7 and T47D cells. To further dissect the regulation of ESR1 transcription by menin, we examined whether and in which way menin could regulate the proximal ESR1 promoter, which has not been fully explored. Using ChIP analysis and reporter gene assays covering - 2500 bp to + 2000 bp of the TSS position, we showed that the activity of the proximal ESR1 promoter was markedly reduced upon menin downregulation independently of H3K4me3 status. Importantly, by analyzing the expression of menin in 354 human breast cancers, we found that a lower expression was associated with ER-negative breast cancer (P = 0.041). Moreover, among the 294 ER-positive breast cancer samples, reduced menin expression was not only associated with larger tumors (P = 0.01) and higher SBR grades (P = 0.005) but also with the luminal B-like breast cancer subtype (P = 0.006). Consistent with our clinical data, we demonstrated that GATA3 and FOXA1, co-factors in ESR1 regulation, interact physically with menin in MCF7 cells, and MEN1 knockdown led to altered protein expression of GATA3, the latter being a known marker of the luminal A subtype, in MCF7 cells. CONCLUSION: Taken together, our data provide clues to the important role of menin in ERα regulation and the formation of breast cancer subtypes.

MEN1 silencing aggravates tumorigenic potential of AR-independent prostate cancer cells through nuclear translocation and activation of JunD and ß-catenin.

BACKGROUND: Recent studies highlighted the increased frequency of AR-low or -negative prostate cancers (PCas) and the importance of AR-independent mechanisms in driving metastatic castration-resistant PCa (mCRPC) development and progression. Several previous studies have highlighted the involvement of the MEN1 gene in PCa. In the current study, we focused on its role specifically in AR-independent PCa cells. METHODS: Cell tumorigenic features were evaluated by proliferation assay, foci formation, colony formation in soft agar, wound healing assay and xenograft experiments in mice. Quantitative RT-PCR, Western blot and immunostaining were performed to determine the expression of different factors in human PCa lines. Different ChIP-qPCR-based assays were carried out to dissect the action of JunD and ß-catenin. RESULTS: We found that MEN1 silencing in AR-independent cell lines, DU145 and PC3, resulted in an increase in anchorage independence and cell migration, accompanied by sustained MYC expression. By searching for factors known to positively regulate MYC expression and play a relevant role in PCa development and progression, we uncovered that MEN1-KD triggered the nuclear translocation of JunD and ß-catenin. ChIP and 3C analyses further demonstrated that MEN1-KD led to, on the one hand, augmented binding of JunD to the MYC 5' enhancer and increased formation of loop structure, and on the other hand, increased binding of ß-catenin to the MYC promoter. Moreover, the expression of several molecular markers of EMT, including E-cadherin, BMI1, Twist1 and HIF-1α, was altered in MEN1-KD DU145 and PC3 cells. In addition, analyses using cultured cells and PC3-GFP xenografts in mice demonstrated that JunD and ß-catenin are necessary for the altered tumorigenic potential triggered by MEN1 inactivation in AR-independent PCa cells. Finally, we observed a significant negative clinical correlation between MEN1 and CTNNB1 mRNA expression in primary PCa and mCRPC datasets. CONCLUSIONS: Our current work highlights an unrecognized oncosuppressive role for menin specifically in AR-independent PCa cells, through the activation of JunD and ß-catenin pathways.

Men1 disruption in Nkx3.1-deficient mice results in ARlow/CD44+ microinvasive carcinoma development with the dysregulated AR pathway.

Dysregulated androgen receptor (AR) plays a crucial role in prostate cancer (PCa) development, though further factors involved in its regulation remain to be identified. Recently, paradoxical results were reported on the implication of the MEN1 gene in PCa. To dissect its role in prostate luminal cells, we generated a mouse model with inducible Men1 disruption in Nkx3.1-deficient mice in which mouse prostatic intraepithelial neoplasia (mPIN) occur. Prostate glands from mutant and control mice were analyzed pathologically and molecularly; cellular and molecular analyses were carried out in PCa cell lines after MEN1 knockdown (KD) by siRNA. Double-mutant mice developed accelerated mPIN and later displayed microinvasive adenocarcinoma. Markedly, early-stage lesions exhibited a decreased expression of AR and its target genes, accompanied by reduced CK18 and E-cadherin expression, suggesting a shift from a luminal to a dedifferentiated epithelial phenotype. Intriguingly, over 60% of menin-deficient cells expressed CD44 at a later stage. Furthermore, MEN1 KD led to the increase in CD44 expression in PC3 cells re-expressing AR. Menin bound to the proximal AR promoter and regulated AR transcription via the H3K4me3 histone mark. Interestingly, the cell proliferation of AR-dependent cells (LNCaP, 22Rv1, and VCaP), but not of AR-independent cells (DU145, PC3), responded strongly to MEN1 silencing. Finally, menin expression was found reduced in some human PCa. These findings highlight the regulation of the AR promoter by menin and the crosstalk between menin and the AR pathway. Our data could be useful for better understanding the increasingly reported AR-negative/NE-negative subtype of PCa and the mechanisms underlying its development.

Gut Microbiome and Metabolome Response of Pu-erh Tea on Metabolism Disorder Induced by Chronic Alcohol Consumption.

This study investigated the protective effects of pu-erh tea extract (PTE) on alcohol-induced microbiomic and metabolomic disorders. In chronic alcohol-exposed mice, PTE ameliorated chronic alcoholic consumption-induced oxidative stress, inflammation, lipid accumulation, and liver and colon damage through modulating microbiomic and metabolomic responses. PTE restored the alcohol-induced fecal microbiota dysbiosis by elevating the relative abundance of potentially beneficial bacteria, for example, Bifidobacterium and Allobaculum, and decreasing the relative abundance of potentially harmful bacteria, for example, Helicobacter and Bacteroides. The alcohol-induced metabolomic disorder was modulated by PTE, which was characterized by regulations of lipid metabolism (sphingolipid, glycerophospholipid, and linoleic acid metabolism), amino acid metabolism (phenylalanine and tryptophan metabolism), and purine metabolism. Besides, the bacterial metabolites of phytochemicals in PTE might contribute to the protective effects of PTE. Overall, PTE could be a functional beverage to treat chronic alcohol consumption-induced microbiomic and metabolomic disorders.

Prebiotic Properties of Green and Dark Tea Contribute to Protective Effects in Chemical-Induced Colitis in Mice: A Fecal Microbiota Transplantation Study.

Green and dark tea extract (GTE/DTE) ameliorate chemical-induced colitis in mice; however, the role of gut microbiota in the anticolitis effects of green and dark tea in mice remains unclear. This study aims to explore the role of modulations in gut microbes mediated by green and dark tea in colitis mice by fecal microbiota transplantation (FMT). Our results indicated that GTE and DTE (5 mg/kg bodyweight/day for 4 weeks) exhibited prebiotic effects on the donor mice. Moreover, the FMT treatments (transferring the microbiota daily from the 1 g/kg bodyweight fecal sample to each recipient) indicated that, compared with the fecal microbiota from the normal diet-treated donor mice, the fecal microbiota from the GTE- and DTE-treated donor mice significantly ameliorate colitis-related symptoms (e.g., loss of bodyweight, colonic inflammation, loss of barrier integrity, and gut microbiota dysbiosis) and downregulated the TLR4/MyD88/NF-κB pathway. Collectively, GTE and DTE ameliorate chemical-induced colitis by modulating gut microbiota.

A fast and simple one-step duplex PCR assay for canine distemper virus (CDV) and canine coronavirus (CCoV) detection.

The one-step polymerase chain reaction (one-step PCR) detection assay is an innovative PCR detection method, eliminating nucleic acid extraction steps, in which samples can be directly added to PCR reagents for testing. For simultaneous detection of CDV and CCoV, a sensitive and specific one-step duplex PCR (one-step dPCR) assay was developed with two pairs of primers that were designed based on H and M gene sequences of CDV and CCoV, respectively. The one-step dPCR with optimized detection conditions has high specificity and sensitivity; independent sequencing assays further verified these results.

Phylogenetic analysis of porcine circovirus type 3 and porcine circovirus type 2 in China detected by duplex nanoparticle-assisted PCR.

Porcine circovirus 2 causes different clinical syndromes resulting in a significant economic loss in the pork industry and PCV3 infection was mainly distributed in the pig herds exhibiting PDNS and reproductive failure symptoms and was widely prevalent in the swine population in several states of the USA. Subsequently, PCV3 was also detected in the Chinese pig herds in Hubei and Shandong Provinces, etc. In a screening of 265 clinical samples from Beijing, Zhejiang, Guangdong and Hebei in China for PCV2, PCV3 and co-infection of PCV2 and PCV3, 14.2%, 16.6% and 6.8% of the samples tested positive, respectively. Under novel and optimized nano-dPCR reaction conditions, two specific fragments of 528 bp (PCV2) and 251 bp (PCV3) were amplified, respectively. No fragments were amplified when other porcine viruses were used as template. In this study, three PCV2 and three PCV3 isolates were evaluated by phylogenetic and alignment analysis and the results revealed that the PCV2 and PCV3 isolates appear new nucleotide substitutions and PCV3 isolates from China still included branch of porcine circovirus in USA.

Isolation and Characterization of Encephalomyocarditis Virus from Dogs in China.

Encephalomyocarditis virus (EMCV) is as a potential zoonotic agent with a wide host range. Here, we describe an EMC virus isolate, identified as EMCV C15, which was successfully obtained from the serum of dogs from animal hospitals. Virus production in cell culture was confirmed by EMCV-specific real-time RT-PCR, indirect immunofluorescence assays and electron microscopy. In addition, the open reading frame sequence (ORF) of the EMCV C15 virus was determined. From sequence comparison and phylogenetic analysis among 24 reference EMCV strains, it appears that the EMCV C15 strain is closely genetically related to strain BEL2887A/91 (>99.0% nucleotide identity). In artificially challenged dogs, the heart and brain were important targets of EMCV C15. This study provides genetic and pathogenic characterization of the EMCV C15 strain isolated in Beijing and calls for sustained surveillance of EMCV infection in China to support better prevention and control of the disease.

A sensitive duplex nanoparticle-assisted PCR assay for identifying porcine epidemic diarrhea virus and porcine transmissible gastroenteritis virus from clinical specimens.

In this study, a novel duplex nanoparticle-assisted polymerase chain reaction (nanoPCR) assay was developed to detect porcine epidemic diarrhea virus (PEDV) and porcine transmissible gastroenteritis virus (TGEV). Two pairs of primers were designed based on the conserved region within the N gene of PEDV and TGEV. In a screening of 114 clinical samples from four provinces in China for PEDV and TGEV, 48.2 and 3.5 % of the samples, respectively, tested positive. Under optimized conditions, the duplex nanoPCR assay had a detection limit of 7.6 × 101 and 8.5 × 101 copies µL-1 for PEDV and TGEV, respectively. The sensitivity of the duplex nanoPCR assay was ten times higher than that of a conventional PCR assay. Moreover, no fragments were amplified when the duplex nanoPCR assay was used to test samples containing other porcine viruses. Our results indicate that the duplex nanoPCR assay described here is useful for the rapid detection of PEDV and TGEV and can be applied in clinical diagnosis.

Concurrent infections of pseudorabies virus and porcine bocavirus in China detected by duplex nanoPCR.

Nanoparticle-assisted polymerase chain reaction (nanoPCR) is a novel method for the simple, rapid, and specific amplification of DNA and has been used to detect viruses. A duplex nanoPCR molecular detection system was developed to detect pseudorabies virus (PRV) and porcine bocavirus (PBoV). Primers were selected to target conserved regions within the PRV gE gene and the PBoV NS1 gene. Under optimized nanoPCR reaction conditions, two specific fragments of 316 bp (PRV) and 996 bp (PBoV) were amplified by the duplex nanoPCR with a detection limit of 6 copies for PRV and 95 copies for PBoV; no fragments were amplified when other porcine viruses were used as template. When used to test 550 clinical samples, the duplex nanoPRC assay and a conventional duplex PCR assay provided very similar results (98.1% consistency); single PRV infections, single PBoV infections, and concurrent PRV and PBoV infections were detected in 37%, 15%, and 9% of the samples, respectively. The results indicate that the novel duplex nanoPCR assay is useful for the rapid detection of PRV and PBoV in pigs.

Development and evaluation of the rVP-ELISA for detection of antibodies against porcine parvovirus.

The gene encoding the VP2 protein of porcine parvovirus (PPV) was expressed in an insect-baculovirus system. The recombinant (r) VP2 was similar antigenically/functionally to the native capsid protein as demonstrated by hemagglutination (HA), Western blotting using PPV positive sera. The purified rVP2 proteins were used as coating antigen to establish a rVP-ELISA method for detection of PPV positive and negative sera from pigs. The optimal operating conditions of the rVP-ELISA were: the concentration of rVP2 proteins coated on the wells was 2 µg/mL; the diluted concentration of serum was 1: 150 and that of the enzyme-labeled antibody was 1: 6000. A total of 596 sera were detected by this assay, and the average positive rate was 87%. Compared with France LSI kit, the result showed that the coincidence rate was 96.7%. In conclusion, the rVP2-ELISA is a sensitive and specific method for detecting antibodies against PPV.