The varicella-zoster virus ORF16 protein promotes both the nuclear transport and the protein abundance of the viral DNA polymerase subunit ORF28.

Although all herpesviruses utilize a highly conserved replication machinery to amplify their viral genomes, different members may have unique strategies to modulate the assembly of their replication components. Herein, we characterize the subcellular localization of seven essential replication proteins of varicella-zoster virus (VZV) and show that several viral replication enzymes such as the DNA polymerase subunit ORF28, when expressed alone, are localized in the cytoplasm. The nuclear import of ORF28 can be mediated by the viral DNA polymerase processivity factor ORF16. Besides, ORF16 could markedly enhance the protein abundance of ORF28. Noteworthily, an ORF16 mutant that is defective in nuclear transport still retained the ability to enhance ORF28 abundance. The low abundance of ORF28 in transfected cells was due to its rapid degradation mediated by the ubiquitin-proteasome system. We additionally reveal that radicicol, an inhibitor of the chaperone Hsp90, could disrupt the interaction between ORF16 and ORF28, thereby affecting the nuclear entry and protein abundance of ORF28. Collectively, our findings imply that the cytoplasmic retention and rapid degradation of ORF28 may be a key regulatory mechanism for VZV to prevent untimely viral DNA replication, and suggest that Hsp90 is required for the interaction between ORF16 and ORF28.

The Impact of Cefuroxime Susceptibility on Aeromonas Necrotizing Fasciitis Outcomes.

Despite aggressive antibiotic therapy and surgical debridement, Aeromonas necrotizing fasciitis (NF) can lead to high amputation and mortality rates. Our study compares the different antibiotic minimum inhibitory concentrations (MICs) via Epsilometer tests (E-tests) between non-survivors and survivors of Aeromonas NF of limbs. A prospective review of 16 patients with Aeromonas NF was conducted for 3.5 years in a tertiary coastal hospital. E-tests were conducted for 15 antimicrobial agents to determine the MIC value for Aeromonas species. These patients were divided into non-survival and survival groups. The clinical outcomes, demographics, comorbidities, presenting signs and symptoms, laboratory findings, and microbiological results between the two periods were compared. A total of four patients died, whereas 12 survived, resulting in a 25% mortality rate. A higher proportion of bloodstream infections (100% vs. 41.7%; p = 0.042), monomicrobial infections (100% vs. 33.3%; p = 0.021), shock (100% vs. 33.3%; p = 0.021), serous bullae (50% vs. 0%; p = 0.009), liver cirrhosis (100% vs. 25%; p = 0.009), chronic kidney disease (100% vs. 33.3%; p = 0.021), lower susceptibility to cefuroxime (25% vs. 83.3%; p = 0.028), and ineffective antibiotic prescriptions (75% vs. 16.7%; p = 0.029) was observed in non-survivors. Aeromonas NF is an extremely rare skin and soft-tissue infection that is associated with high mortality, bacteremia, antibiotic resistance, and polymicrobial infection. Therefore, antibiotic regimen selection is rendered very challenging. To improve clinical outcomes and irrational antimicrobial usage, experienced microbiologists can help physicians identify specific pathogens and test MIC.

Interaction and assembly of the DNA replication core proteins of Kaposi's sarcoma-associated herpesvirus.

IMPORTANCE: Eukaryotic DNA replication is a highly regulated process that requires multiple replication enzymes assembled onto DNA replication origins. Due to the complexity of the cell's DNA replication machinery, most of what we know about cellular DNA replication has come from the study of viral systems. Herein, we focus our study on the assembly of the Kaposi's sarcoma-associated herpesvirus core replication complex and propose a pairwise protein-protein interaction network of six highly conserved viral core replication proteins. A detailed understanding of the interaction and assembly of the viral core replication proteins may provide opportunities to develop new strategies against viral propagation.

Comparative Study of the Cytokine Profiles of Serum and Tissues from Patients with the Ossification of the Posterior Longitudinal Ligament.

BACKGROUND: The ossification of the posterior longitudinal ligament (OPLL) is one of the contributing factors leading to severe cervical spondylotic myelopathy (CSM). The mechanism causing ossification is still unclear. The current study was designed to analyze the specimens of patients with or without OPLL. METHODS: The study collected 51 patients with cervical spondylosis. There were six serum samples in both the non-OPLL (NOPLL) and OPLL groups. For tissue analysis, there were seven samples in the NOPLL group and five samples in the OPLL group. The specimens of serum and tissue were analyzed by using Human Cytokine Antibody Arrays to differentiate biomarkers between the OPLL and NOPLL groups, as well as between serum and OPLL tissue. Immunohistochemical staining of the ligament tissue was undertaken for both groups. RESULTS: For OPLL vs. NOPLL, the serum leptin levels are higher in the OPLL group, corroborating others' observations that it may serve as a disease marker. In the tissue, angiogenin (ANG), osteopontin (OPN), and osteopro-tegerin (OPG) are higher than they are in the OPLL group (p < 0.05). For serum vs. OPLL tissue, many chemotactic cytokines demonstrated elevated levels of MIP1 delta, MCP-1, and RANTES in the serum, while many cytokines promoting or regulating bone genesis were up-regulated in tissue (oncostatin M, FGF-9, LIF, osteopontin, osteoprotegerin, TGF-beta2), as well as the factor that inhibits osteoclastogenesis (IL-10), with very few cytokines responsible for osteoclastogenesis. Molecules promoting angiogenesis, including angiotensin, vEGF, and osteoprotegerin, are abundant in the OPLL tissue, which paves the way for robust bone growth.

Cytokine-Induced Myeloid-Derived Suppressor Cells Demonstrate Their Immunoregulatory Functions to Prolong the Survival of Diabetic Mice.

Type 1 diabetes is an inflammatory state. Myeloid-derived suppressive cells (MDSCs) originate from immature myeloid cells and quickly expand to control host immunity during infection, inflammation, trauma, and cancer. This study presents an ex vivo procedure to develop MDSCs from bone marrow cells propagated from granulocyte-macrophage-colony-stimulating factor (GM-CSF), interleukin (IL)-6, and IL-1ß cytokines expressing immature morphology and high immunosuppression of T-cell proliferation. The adoptive transfer of cytokine-induced MDSCs (cMDSCs) improved the hyperglycemic state and prolonged the diabetes-free survival of nonobese diabetic (NOD) mice with severe combined immune deficiency (SCID) induced by reactive splenic T cells harvested from NOD mice. In addition, the application of cMDSCs reduced fibronectin production in the renal glomeruli and improved renal function and proteinuria in diabetic mice. Moreover, cMDSCs use mitigated pancreatic insulitis to restore insulin production and reduce the levels of HbA1c. In conclusion, administering cMDSCs propagated from GM-CSF, IL-6, and IL-1ß cytokines provides an alternative immunotherapy protocol for treating diabetic pancreatic insulitis and renal nephropathy.

Propess versus prostin for induction of labour in term primiparous women.

BACKGROUND: The rate of induction of labour has increased over the decades and numerous medications are available in the market. This study compares the efficacy and safety between dinoprostone slow-release pessary (Propess) and dinoprostone tablet (Prostin) for labour induction at term in nulliparous women. METHODS: This was a prospective single-blind randomized controlled trial conducted in a tertiary medical centre in Taiwan from September 1, 2020 to February 28, 2021. We recruited nulliparous women at term with a singleton pregnancy, fetus in cephalic presentation, an unfavourable cervix, and the cervical length had been measured by transvaginal sonography three times during labour induction. The main outcomes are duration from induction of labour to vaginal delivery, vaginal delivery rate, maternal and neonatal complication rates. RESULTS: In both groups, Prostin and Propess, 30 pregnant women were enrolled. The Propess group had higher vaginal delivery rate but it did not meet statistically significant difference. The Prostin group had significantly higher rate of adding oxytocin for augmentation (p = 0.0002). No significant difference was observed in either labouring course, maternal or neonatal outcomes. The probability of vaginal delivery was independently related to the cervical length measured by transvaginal sonography 8 h after Prostin or Propess administration as well as neonatal birth weight. CONCLUSION: Both Prostin and Propess can be used as cervical ripening agents with similar efficacy and without significant morbidity. Propess administration was associated with higher vaginal delivery rate and less need to add oxytocin. Intrapartum measurement of cervical length is helpful in predicting successful vaginal delivery.

Characterization of Human Norovirus Nonstructural Protein NS1.2 Involved in the Induction of the Filamentous Endoplasmic Reticulum, Enlarged Lipid Droplets, LC3 Recruitment, and Interaction with NTPase and NS4.

Human noroviruses (HuNVs) are the leading cause of gastroenteritis worldwide. NS1.2 is critical for HuNV pathogenesis, but the function is still unclear. The GII NS1.2 of HuNVs, unlike GI NS1.2, was localized to the endoplasmic reticulum (ER) and lipid droplets (LDs) and is accompanied by a distorted-filamentous ER morphology and aggregated-enlarged LDs. LC3 was recruited to the NS1.2-localized membrane through an autophagy-independent pathway. NS1.2, expressed from a cDNA clone of GII.4 norovirus, formed complexes with NTPase and NS4, which exhibited aggregated vesicle-like structures that were also colocalized with LC3 and LDs. NS1.2 is structurally divided into three domains from the N terminus: an inherently disordered region (IDR), a region that contains a putative hydrolase with the H-box/NC catalytic center (H-box/NC), and a C-terminal 251-330 a.a. region containing membrane-targeting domain. All three functional domains of NS1.2 were required for the induction of the filamentous ER. The IDR was essential for LC3 recruitment by NS1.2. Both the H-Box/NC and membrane-targeting domains are required for the induction of aggregated-enlarged LDs, NS1.2 self-assembly, and interaction with NTPase. The membrane-targeting domain was sufficient to interact with NS4. The study characterized the NS1.2 domain required for membrane targeting and protein-protein interactions, which are crucial for forming a viral replication complex.

Rational Use of Antibiotics and Education Improved Aeromonas Necrotizing Fasciitis Outcomes in Taiwan: A 19-Year Experience.

BACKGROUND: Aeromonas necrotizing fasciitis (NF) causes high rates of amputation and mortality, even after aggressive surgical debridement and antibacterial therapy. This study investigated the effects of rational use of antibiotics and education by infectious disease (ID) physicians on Aeromonas NF treatment outcomes. METHODS: Retrospective review for conducted for four years (period I, without an ID physician, December 2001 to December 2005) and 15 years (period II, with an ID physician, January 2006 to March 2021). In period II, the hospital-wide computerized antimicrobial approval system (HCAAS) was also implemented. A pretest-posttest time series analysis compared the two periods. Differences in clinical outcomes, demographics, comorbidities, signs and symptoms, laboratory findings, Aeromonas antibiotic susceptibility, and antibiotic regimens were compared between the two periods. RESULTS: There were 19 patients in period I and 53 patients in period II. Patients had a lower rate of amputation or mortality in period II (35.8%) compared with period I (63.2%). Forty-four patients (61.1%) had polymicrobial infections. In the emergency room, the rate of misdiagnosis decreased from 47.4% in period I to 28.3% in period II, while effective empiric antibiotic usage increased from 21.1% in period I to 66.0% in period II. After the ID physician's adjustment, 69.4% received monotherapy in period II compared to 33.3% in period I. CONCLUSIONS: Because Aeromonas NF had a high mortality rate and was often polymicrobial, choosing an antibiotic regimen was difficult. Using the HCAAS by an experienced ID physician can improve rational antibiotic usage and clinical outcomes in Aeromonas NF.

Exogenous FGF-1 Differently Regulates Oligodendrocyte Replenishment in an SCI Repair Model and Cultured Cells.

We studied the phenotypes in an oligodendrocyte genesis site at the acute stage of spinal cord injury, when we observed regenerated ascending neurites. Pan-oligodendrocyte marker OLIG2+ cells were more in fibroblast growth factor (FGF)-1-treated rats (F group) than in non-treated (T group) in this site, while the number of NG2+OX42- oligodendrocyte progenitor cell (OPC), CNPase+ OPC, Nkx2.2+ OPC, and APC+ remyelinating oligodendrocytes was less in the F group. Paradoxically, when we label the rats with pulsed bromodeoxyuridine (BrdU), we found that the mitotic NKX2.2+ OPC cells are more in the F group than in the T group. We tested the embryonic spinal cord mixed culture. FGF treatment resulted in more NG2(+) CNPase (+) than non-FGF-1-treated culture, while the more mature NG2(-) CNPase(+) cell numbers were reduced. When we block the FGF receptor in the injured rat model, the NG2+OX42- cell numbers were increased to be comparable to non-FGF-1 rats, while this failed to bring back the APC+ mature oligodendrocyte cell numbers. As migration of OPC toward injury is a major factor that was absent from the cell culture, we tested 8 mm away from the injury center, and found there were more NG2+ cells with FGF-1 treatment. We proposed that it was possibly a combination of migration and proliferation that resulted in a reduction in the NG2+ OPC population at the oligodendrocyte genesis site when FGF-1 was added to the spinal cord injury in vivo.

Kaposi's Sarcoma-Associated Herpesvirus ORF50 Protein Represses Cellular MDM2 Expression via Suppressing the Sp1- and p53-Mediated Transactivation.

The Kaposi's sarcoma-associated herpesvirus (KSHV)-encoded ORF50 protein is a potent transcriptional activator essential for triggering KSHV lytic reactivation. Despite extensive studies, little is known about whether ORF50 possesses the ability to repress gene expression or has an antagonistic action to cellular transcription factors. Previously, we demonstrated that human oncoprotein MDM2 can promote the degradation of ORF50 protein. Herein, we show that abundant ORF50 expression in cells can conversely downregulate MDM2 expression via repressing both the upstream (P1) and internal (P2) promoters of the MDM2 gene. Deletion analysis of the MDM2 P1 promoter revealed that there were two ORF50-dependent negative response elements located from -102 to -63 and from -39 to +1, which contain Sp1-binding sites. For the MDM2 P2 promoter, the ORF50-dependent negative response element was identified in the region from -110 to -25, which is coincident with the location of two known p53-binding sites. Importantly, we further demonstrated that overexpression of Sp1 or p53 in cells indeed upregulated MDM2 expression; however, coexpression with ORF50 protein remarkably reduced the Sp1- or p53-mediated MDM2 upregulation. Collectively, our findings propose a reciprocal negative regulation between ORF50 and MDM2 and uncover that ORF50 decreases MDM2 expression through repressing Sp1- and p53-mediated transactivation.

Caspase-Mediated Cleavage of the Transcription Factor Sp3: Possible Relevance to Cancer and the Lytic Cycle of Kaposi's Sarcoma-Associated Herpesvirus.

The open reading frame 50 (ORF50) protein of Kaposi's sarcoma-associated herpesvirus (KSHV) is the master regulator essential for initiating the viral lytic cycle. Previously, we have demonstrated that the ORF50 protein can cooperate with Sp3 to synergistically activate a set of viral and cellular gene promoters through highly conserved ORF50-responsive elements that harbor a Sp3-binding motif. Herein, we show that Sp3 undergoes proteolytic cleavage during the viral lytic cycle, and the cleavage of Sp3 is dependent on caspase activation. Since similar cleavage patterns of Sp3 could be detected in both KSHV-positive and KSHV-negative lymphoma cells undergoing apoptosis, the proteolytic cleavage of Sp3 could be a common event during apoptosis. Mutational analysis identifies 12 caspase cleavage sites in Sp3, which are situated at the aspartate (D) positions D17, D19, D180, D273, D275, D293, D304 (or D307), D326, D344, D530, D543, and D565. Importantly, we noticed that three stable Sp3 C-terminal fragments generated through cleavage at D530, D543, or D565 encompass an intact DNA-binding domain. Like the full-length Sp3, the C-terminal fragments of Sp3 could still retain the ability to cooperate with ORF50 protein to activate specific viral and cellular gene promoters synergistically. Collectively, our findings suggest that despite the proteolytic cleavage of Sp3 under apoptotic conditions, the resultant Sp3 fragments may retain biological activities important for the viral lytic cycle or for cellular apoptosis. IMPORTANCE The ORF50 protein of Kaposi's sarcoma-associated herpesvirus (KSHV) is the key viral protein that controls the switch from latency to lytic reactivation. It is a potent transactivator that can activate target gene promoters via interacting with other cellular DNA-binding transcription factors, such as Sp3. In this report, we show that Sp3 is proteolytically cleaved during the viral lytic cycle, and up to 12 caspase cleavage sites are identified in Sp3. Despite the proteolytic cleavage of Sp3, several resulting C-terminal fragments that have intact zinc-finger DNA-binding domains still retain substantial influence in the synergy with ORF50 to activate specific gene promoters. Overall, our studies elucidate the caspase-mediated cleavage of Sp3 and uncover how ORF50 utilizes the cleavage fragments of Sp3 to transactivate specific viral and cellular gene promoters.

The Prenatal Diagnosis and Clinical Outcomes of Fetuses With 15q11.2 Copy Number Variants: A Case Series of 36 Patients.

Prenatal genetic counseling of fetuses diagnosed with 15q11.2 copy number variants (CNVs) involving the BP1-BP2 region is difficult due to limited information and controversial opinion on prognosis. In total, we collected the data of 36 pregnant women who underwent prenatal microarray analysis from 2010 to 2017 and were assessed at National Taiwan University Hospital. Comparison of the maternal characteristics, prenatal ultrasound findings, and postnatal outcomes among the different cases involving the 15q11.2 BP1-BP2 region were presented. Out of the 36 fetuses diagnosed with CNVs involving the BP1-BP2 region, five were diagnosed with microduplications and 31 with microdeletions. Among the participants, 10 pregnant women received termination of pregnancy and 26 gave birth to healthy individuals (27 babies in total). The prognoses of 15q11.2 CNVs were controversial and recent studies have revealed its low pathogenicity. In our study, the prenatal abnormal ultrasound findings were recorded in 12 participants and were associated with 15q11.2 deletions. No obvious developmental delay or neurological disorders were detected in early childhood.

Effect of Xanthium Strumarium on HIV-1 5'-LTR Transcriptional Activity and Viral Reactivation in Latently Infected Cells.

Chinese herbal medicines (CHMs) are widely used in Asian countries. They show multiple pharmacological activities, including antiviral activities. The 5'-long terminal repeat (LTR) region of HIV-1, required for viral transcription, is a potential drug target for HIV-1 reactivation and intrinsic cell death induction of infected or latently infected cells. Modulation of HIV-1 reactivation requires interactions between host cell proteins and viral 5'-LTR elements. By evaluation of two CHMs- Xanthium strumarium and Pueraria montana, we found that 1) X. strumarium reactivated HIV-1 latently infected cells in J-Lat 8.4, J-Lat 9.2, U1, and ACH-2 cells in vitro; 2) 27 nuclear regulatory proteins were associated with HIV-1 5'-LTR using deoxyribonucleic acid affinity pull-down and LC-MS/MS analyses; and 3) among them, silencing of XRCC6 reactivated HIV-1 5'-LTR transcriptional activity. We found that X. strumarium inhibits the 5'-LTR associated XRCC6 nuclear regulatory proteins, increases its viral 5'-LTR promoter transcriptional activity, and reactivates HIV-1 latently infected cells in vitro. These findings may contribute to understanding the 5'-LTR activity and the host cell nuclear regulatory protein machinery for reactivating HIV-1 and for future investigations to eradicate and cure HIV-1 infection.

Identification and Characterization of Human Norovirus NTPase Regions Required for Lipid Droplet Localization, Cellular Apoptosis, and Interaction with the Viral P22 Protein.

The human norovirus (HuNV)-encoded nucleoside-triphosphatase (NTPase) is a multifunctional protein critically involved in viral replication and pathogenesis. Previously, we have shown that the viral NTPase is capable of forming vesicle clusters in cells, interacting with other viral proteins such as P22, and promoting cellular apoptosis. Herein, we demonstrate that NTPase-associated vesicle clusters correspond to lipid droplets (LDs) wrapped by the viral protein and show that NTPase-induced apoptosis is mediated through both caspase-8- and caspase-9-dependent pathways. Deletion analysis revealed that the N-terminal 179-amino-acid (aa) region of NTPase encompasses two LD-targeting motifs (designated LTM-1 and LTM-2), two apoptosis-inducing motifs, and multiple regulatory regions. Interestingly, the identified LTM-1 and LTM-2, which are located from aa 1 to 50 and from aa 51 to 90, respectively, overlap with the two apoptosis-inducing motifs. Although there was no positive correlation between the extent of LD localization and the degree of cellular apoptosis for NTPase mutants, we noticed that mutant proteins defective in LD-targeting ability could not induce cellular apoptosis. In addition to LD targeting, the amphipathic LTM-1 and LTM-2 motifs could have the potential to direct fusion proteins to the endoplasmic reticulum (ER). Furthermore, we found that the LTM-1 motif is a P22-interacting motif. However, P22 functionally augmented the proapoptotic activity of the LTM-2 fusion protein but not the LTM-1 fusion protein. Overall, our findings propose that NTPase may participate in multiple cellular processes through binding to LDs or to the ER via its N-terminal amphipathic helix motifs. IMPORTANCE Human noroviruses (HuNVs) are the major agent of global gastroenteritis outbreaks. However, due to the lack of an efficient cell culture system for HuNV propagation, functions of the viral-encoded proteins in host cells are still poorly understood. In the current study, we present that the viral NTPase is a lipid droplet (LD)-associated protein, and we identify two LD-targeting motifs, LTM-1 and LTM-2, in its N-terminal domain. In particular, the identified LTM-1 and LTM-2 motifs, which contain a hydrophobic region and an amphipathic helix, are also capable of delivering the fusion protein to the endoplasmic reticulum (ER), promoting cellular apoptosis, and physically or functionally associating with another viral protein P22. Since LDs and the ER have been linked to several biological functions in cells, our study therefore proposes that the norovirus NTPase may utilize LDs or the ER as replication platforms to benefit viral replication and pathogenesis.

Rapid Trio Exome Sequencing for Autosomal Recessive Renal Tubular Dysgenesis in Recurrent Oligohydramnios.

Oligohydramnios is not a rare prenatal finding. However, recurrent oligohydramnios is uncommon, and genetic etiology should be taken into consideration. We present two families with recurrent fetal oligohydramnios that did not respond to amnioinfusion. Rapid trio-whole-exome sequencing (WES) revealed mutations in the AGT gene in both families within 1 week. The first family had a compound heterozygous mutation with c.856 + 1G > T and c.857-619_1269 + 243delinsTTGCCTTGC changes. The second family had homozygous c.857-619_1269 + 243delinsTTGCCTTGC mutations. AGT gene mutation may lead to autosomal recessive renal tubular dysgenesis, a rare and lethal disorder that can result in early neonatal death. Both the alleles identified are known alleles associated with pathogenicity. Our findings suggest that trio-WES analysis may help rapidly identify causative etiologies that can inform prompt counseling and decision-making prenatally.

The Epstein-Barr Virus Lytic Protein BMLF1 Induces Upregulation of GRP78 Expression through ATF6 Activation.

The unfolded protein response (UPR) is an intracellular signaling pathway essential for alleviating the endoplasmic reticulum (ER) stress. To support the productive infection, many viruses are known to use different strategies to manipulate the UPR signaling network. However, it remains largely unclear whether the UPR signaling pathways are modulated in the lytic cycle of Epstein-Barr virus (EBV), a widely distributed human pathogen. Herein, we show that the expression of GRP78, a central UPR regulator, is up-regulated during the EBV lytic cycle. Our data further revealed that knockdown of GRP78 in EBV-infected cell lines did not substantially affect lytic gene expression; however, GRP78 knockdown in these cells markedly reduced the production of virus particles. Importantly, we identified that the early lytic protein BMLF1 is the key regulator critically contributing to the activation of the grp78 gene promoter. Mechanistically, we found that BMLF1 can trigger the proteolytic cleavage and activation of the UPR senor ATF6, which then transcriptionally activates the grp78 promoter through the ER stress response elements. Our findings therefore provide evidence for the connection between the EBV lytic cycle and the UPR, and implicate that the BMLF1-mediated ATF6 activation may play critical roles in EBV lytic replication.

Different types of bullae of limbs with necrotizing fasciitis predict different outcome: a prospective study.

STUDY OBJECTIVE: Necrotizing fasciitis (NF) is an uncommon life-threatening necrotizing skin and soft tissue infection. Bullae are special skin manifestations of NF. This study was conducted to analyze the differences between different types of bullae of limbs with NF for providing the information to emergency treatment. METHODS: From April 2015 to August 2018, patients were initially enrolled based on surgical confirmation of limbs with NF. According to the presence of different bullae types, patients were divided into no bullae group (Group N), serous-filled bullae group (Group S), and hemorrhagic bullae group (Group H). Data such as demographics, clinical outcomes, microbiological results, presenting symptoms/signs, and laboratory findings were compared among these groups. RESULTS: In total, 187 patients were collected, with 111 (59.4%) patients in Group N, 35 (18.7%) in Group S, and 41 (21.9%) in Group H. Group H had the highest incidence of amputation, required intensive care unit care, and most patients infected with Vibrio species. In Group N, more patients were infected with Staphylococcus spp. than Group H. In Group S, more patients were infected with ß-hemolytic Streptococcus than Group H. Patients with bacteremia, shock, skin necrosis, anemia, and longer prothrombin time constituted higher proportions in Group H and S than in Group N. CONCLUSIONS: In southern Taiwan, patients with NF accompanied by hemorrhagic bullae appear to have more bacteremia, Vibrio infection, septic shock, and risk for amputation. If the physicians at the emergency department can detect for the early signs of NF as soon as possible, and more patient's life and limbs may be saved.

Association Between Preoperative Blood Glucose Level and Hospital Length of Stay for Patients Undergoing Appendectomy or Laparoscopic Cholecystectomy.

OBJECTIVE: To evaluate the effect of preoperative blood glucose (POBG) level on hospital length of stay (LOS) in patients undergoing appendectomy or laparoscopic cholecystectomy. RESEARCH DESIGN AND METHODS: We conducted a retrospective cohort study of patients aged ≥18 years who had undergone appendectomy or laparoscopic cholecystectomy procedures between 2005 and 2016 at a tertiary medical center in Taiwan. The association between POBG level and LOS was evaluated using a multivariable quasi-Poisson regression with robust variance. Multiple imputations were performed to replace missing values. RESULTS: We included 8,291 patients; 4,025 patients underwent appendectomy (appendectomy group) and 4,266 underwent laparoscopic cholecystectomy (laparoscopic cholecystectomy group). In the appendectomy group, patients with POBG levels of ≥123 mg/dL (adjusted relative risk [aRR] 1.19; 95% CI 1.06-1.33) had a 19% higher risk of having a LOS of >3 days than did those with POBG levels of <106 mg/dL. In the laparoscopic cholecystectomy group, patients with POBG levels of ≥128 mg/dL also had a significantly higher risk of having a LOS of >3 days (aRR 1.17; 95% CI 1.07-1.29) than did those with POBG levels of <102 mg/dL. A positive dose-response curve between POBG and an adjusted risk of a LOS of >3 days was observed, although the curve starts to flatten at a POBG level of ∼130 mg/dL. CONCLUSIONS: We demonstrated that a higher POBG level was significantly associated with a prolonged LOS for patients undergoing appendectomy or laparoscopic cholecystectomy. The optimal POBG level may be lower than that commonly perceived.

Virtual Screening and In Vitro Evaluation of PD-1 Dimer Stabilizers for Uncoupling PD-1/PD-L1 Interaction from Natural Products.

Genetic mutations accumulated overtime could generate many growth and survival advantages for cancer cells, but these mutations also mark cancer cells as targets to be eliminated by the immune system. To evade immune surveillance, cancer cells adopted different intrinsic molecules to suppress immune response. PD-L1 is frequently overexpressed in many cancer cells, and its engagement with PD-1 on T cells diminishes the extent of cytotoxicity from the immune system. To resume immunity for fighting cancer, several therapeutic antibodies disrupting the PD-1/PD-L1 interaction have been introduced in clinical practice. However, their immunogenicity, low tissue penetrance, and high production costs rendered these antibodies beneficial to only a limited number of patients. PD-L1 dimer formation shields the interaction interface for PD-1 binding; hence, screening for small molecule compounds stabilizing the PD-L1 dimer may make immune therapy more effective and widely affordable. In the current study, 111 candidates were selected from over 180,000 natural compound structures through virtual screening, contact fingerprint analysis, and pharmacological property prediction. Twenty-two representative candidates were further evaluated in vitro. Two compounds were found capable of inhibiting the PD-1/PD-L1 interaction and promoting PD-L1 dimer formation. Further structure optimization and clinical development of these lead inhibitors will eventually lead to more effective and affordable immunotherapeutic drugs for cancer patients.

Sp3 Transcription Factor Cooperates with the Kaposi's Sarcoma-Associated Herpesvirus ORF50 Protein To Synergistically Activate Specific Viral and Cellular Gene Promoters.

The Kaposi's sarcoma-associated herpesvirus (KSHV)-encoded open reading frame 50 (ORF50) protein is the key transactivator responsible for the latent-to-lytic switch. Here, we investigated the transcriptional activation of the ORF56 gene (encoding a primase protein) by ORF50 and successfully identified an ORF50-responsive element located in the promoter region between positions -97 and -44 (designated 56p-RE). This 56p-RE element contains a noncanonical RBP-Jκ-binding sequence and a nonconsensus Sp1/Sp3-binding sequence. Electrophoretic mobility shift assays revealed that RBP-Jκ, Sp3, and ORF50 could form stable complexes on the 56p-RE element. Importantly, transient-reporter analysis showed that Sp3, but not RBP-Jκ or Sp1, acts in synergy with ORF50 to activate the 56p-RE-containing reporter construct, and the synergy mainly depends on the Sp1/Sp3-binding region of the 56p-RE element. Sequence similarity searches revealed that the promoters for ORF21 (thymidine kinase), ORF60 (ribonucleotide reductase, small subunit), and cellular interleukin-10 (IL-10) contain a sequence motif similar to the Sp1/Sp3-binding region of the 56p-RE element, and we found that these promoters could also be synergistically activated by ORF50 and Sp3 via the conserved motifs. Noteworthily, the conversion of the Sp1/Sp3-binding sequence of the 56p-RE element into a consensus high-affinity Sp-binding sequence completely lost the synergistic response to ORF50 and Sp3. Moreover, transcriptional synergy could not be detected through other ORF50-responsive elements from the viral PAN, K12, ORF57, and K6 promoters. Collectively, the results of our study demonstrate that ORF50 and Sp3 can act in synergy on the transcription of specific gene promoters, and we find a novel conserved cis-acting motif in these promoters essential for transcriptional synergy.IMPORTANCE Despite the critical role of ORF50 in the KSHV latent-to-lytic switch, the molecular mechanism by which ORF50 activates its downstream target genes, especially those that encode the viral DNA replication enzymes, is not yet fully understood. Here, we find that ORF50 can cooperate with Sp3 to synergistically activate promoters of the viral ORF56 (primase), ORF21 (thymidine kinase), and ORF60 (ribonucleotide reductase) genes via similar Sp1/Sp3-binding motifs. Additionally, the same synergistic effect can be seen on the promoter of the cellular IL-10 gene. Overall, our data reveal an important role for Sp3 in ORF50-mediated transactivation, and we propose a new subclass of ORF50-responsive elements in viral and cellular promoters.