Characterization of a pangolin SARS-CoV-2-related virus isolate that uses the human ACE2 receptor.

Various SARS-CoV-2-related coronaviruses have been increasingly identified in pangolins, showing a potential threat to humans. Here we report the infectivity and pathogenicity of the SARS-CoV-2-related virus, PCoV-GX/P2V, which was isolated from a Malayan pangolin (Manis javanica). PCoV-GX/P2V could grow in human hepatoma, colorectal adenocarcinoma cells, and human primary nasal epithelial cells. It replicated more efficiently in cells expressing human angiotensin-converting enzyme 2 (hACE2) as SARS-CoV-2 did. After intranasal inoculation to the hACE2-transgenic mice, PCoV-GX/P2V not only replicated in nasal turbinate and lungs, but also caused interstitial pneumonia, characterized by infiltration of mixed inflammatory cells and multifocal alveolar hemorrhage. Existing population immunity established by SARS-CoV-2 infection and vaccination may not protect people from PCoV-GX/P2V infection. These findings further verify the hACE2 utility of PCoV-GX/P2V by in vivo experiments using authentic viruses and highlight the importance for intensive surveillance to prevent possible cross-species transmission.

Isolation and characterization of a pangolin-borne HKU4-related coronavirus that potentially infects human-DPP4-transgenic mice.

We recently detected a HKU4-related coronavirus in subgenus Merbecovirus (named pangolin-CoV-HKU4-P251T) from a Malayan pangolin1. Here we report isolation and characterization of pangolin-CoV-HKU4-P251T, the genome sequence of which is closest to that of a coronavirus from the greater bamboo bat (Tylonycteris robustula) in Yunnan Province, China, with a 94.3% nucleotide identity. Pangolin-CoV-HKU4-P251T is able to infect human cell lines, and replicates more efficiently in cells that express human-dipeptidyl-peptidase-4 (hDPP4)-expressing and pangolin-DPP4-expressing cells than in bat-DPP4-expressing cells. After intranasal inoculation with pangolin-CoV-HKU4-P251, hDPP4-transgenic female mice are likely infected, showing persistent viral RNA copy numbers in the lungs. Progressive interstitial pneumonia developed in the infected mice, characterized by the accumulation of macrophages, and increase of antiviral cytokines, proinflammatory cytokines, and chemokines in lung tissues. These findings suggest that the pangolin-borne HKU4-related coronavirus has a potential for emerging as a human pathogen by using hDPP4.

Radiographic patterns and characteristics of sagittal profiles in normal spinopelvic curvatures: An explicit depiction of the distribution of lumbar vertebral bodies and discs.

The current study aimed to investigate the sagittal morphology of the spinopelvic complex and the components of the lumbar spine in the normal population. In total, 132 adult volunteers were retrospectively evaluated and divided into four groups according to the Roussouly classification. Statistical analysis of radiological parameters, including lumbar lordosis (LL), thoracic kyphosis (TK), pelvic incidence (PI), pelvic tilt (PT), sacral slope (SS), PI-LL, LL-TK, lumbar vertebral lordosis from L1 to L5 (L1L-L5L), the intervertebral angle from L1-L2 to L5-S1 (IVA1-2-IVA5-1), segmental lordosis from L1 to L5 (S1L-S5L), the proportion of L1-L5 (L1%-L5%), the proportion of the intervertebral angle from L1-L2 to L5-S1 (IVA1-2%-IVA5-1%), and proportion of segmental lordosis from L1 to L5 (S1L%-S5L%), was performed. Based on the classification, type II (n = 46) was the most common, followed by type I (n = 39), type III (n = 36), and type IV (n = 11). The quantitative values of the sagittal parameters of the four groups were obtained. Results showed a significant difference in terms of LL, PI, SS, and LL-TK. Further, L1%, L2%, L3%, IVA1-2%, IVA2-3%, S1L%, S2L%, and S3L% had an increasing trend. PI was positively correlated with LL, S1L, S2L, S3L, S4L, S1L%, and S2L%, but not with S5L%. In conclusion, pelvic parameters had a significant effect on lumbar curvature and lordosis distribution. Further, the abovementioned results were beneficial for individual surgical decision-making regarding targeted intervertebral angle, screw-insertion dimension, and rod contouring.

Diffuse Idiopathic Skeletal Hyperostosis Combined with Thoracic Spinal Stenosis Treated by Endoscopic Surgery: A Case Report.

CASE: A 48-year-old man presented to our facility with debilitating motor and sensory symptoms due to advanced T10-11 thoracic spinal stenosis secondary to diffuse idiopathic skeletal hyperostosis (DISH). The patient's condition was addressed with endoscopic spine surgery through a yet-to-be-reported interlaminar approach, and at the 12-month follow-up, his neurologic function was significantly improved. CONCLUSION: Select patients with symptomatic thoracic spinal stenosis secondary to DISH can be effectively managed with endoscopic spine surgery through an interlaminar approach by clinicians with extensive endoscopic spine experience.

Outcomes and prognostic factors for patients with cervical esophageal cancer undergoing definitive radiotherapy or chemoradiotherapy.

Cervical esophageal cancer (CEC) is uncommon, accounting for less than 5% of all esophageal cancers. The management of CEC is controversial. This study investigated treatment outcomes and prognostic factors of survival in CEC patients undergoing definitive radiotherapy or concurrent chemoradiotherapy (CCRT). Ninety-one CEC patients were treated by intensity-modulated radiation therapy (IMRT) and three-dimensional conformal radiation therapy (3DCRT) between July 2007 and September 2017. The mean prescription dose was 64 Gy (range 54-70 Gy) delivered as 1.8-2.2 Gy per fraction per day, 5 days a week. Out of 91 patients, 34 received concurrent cisplatin-based chemotherapy (CT) including 18 patients who also received neoadjuvant CT. Overall survival (OS), locoregional failure-free survival (LRFFS), and progression-free survival (PFS) were estimated by the Kaplan-Meier method. Prognostic factors of survival were determined in univariate (log-rank test) and multivariate (Cox proportional hazard model) analysis. Treatment-related toxicity was also assessed. Median follow-up time for all patients was 19 months. Two-year OS, LRFFS and PFS of all patients were 58.2%, 52.5% and 48.1%, respectively. Clinical stage was an independent prognostic factor for OS (HR = 2.35, 95% CI: 1.03-5.37, p = 0.042), LRFFS (HR = 3.84, 95% CI: 1.38-10.69, p = 0.011), and PFS (HR = 2.68, 95% CI: 1.11-6.45, p = 0.028). Hoarseness was an independent prognostic factor for OS (HR = 2.10, 95% CI: 1.05-4.19, p = 0.036). CCRT was independently associated with better LRFFS (HR = 0.33, 95% CI: 0.14-0.79, p = 0.012). 3DCRT and IMRT with concurrent CT is well-tolerated and may improve local tumor control in CEC patients. Advanced clinical stage and hoarseness are adverse prognostic factors for OS, LRFFS, and PFS in CEC.

Paeoniflorin Inhibits Migration- and Invasion-Promoting Capacities of Gastric Cancer Associated Fibroblasts.

OBJECTIVE: To investigate the inhibitory effects of paeoniflorin on migration- and invasion-promoting capacities of gastric cancer associated fibroblasts (GCAFs) and to explore the molecular mechanism underlying the effects. METHODS: Paired gastric normal fifbroblast (GNF) and GCAF cultures were established from resected tissues. GCAFs were treated with control medium, or 2.5, 5 or 10 µg/mL paeoniflorin. Conditioned media were prepared from GNFs, GCAFs, control-treated GCAFs and paeoniflorin-treated GCAFs, and used to culture AGS human gastric cancer cells. The migration and invasion capacities of AGS cells were determined with wound healing test and transwell invasion assay, respectively. The interleukin 6 (IL-6) mRNA and microRNA-149 expression in GCAFs were detected by reverse transcription-quantitative polymerase chain reaction. The IL-6 protein expression and secretion by GCAFs were measured with Western blot and enzyme-linked immunosorbent assay analysis, respectively. The protein levels of phosphorylated signal transducer and activator of transcription 3 (STAT3), matrix metalloproteinase (MMP) and MMP9 in AGS cells were examined by Western blot. RESULTS: GCAFs displayed enhanced capacities to induce AGS cell migration and invasion as compared with GNFs. Paeoniflorin treatment significantly inhibited the migration- and invasion-promoting capacities of GCAFs (P<0.05). GCAFs produced and secreted more IL-6 into the conditioned medium than GNFs, leading to over-activation of STAT3-MMP signaling in AGS cells. Paeoniflorin suppressed IL-6 production and secretion by up-regulating microRNA149 expression in GCAFs, and subsequently prevented GCAFs from activating IL-6-STAT3-MMP signaling of AGS cells. CONCLUSIONS: Paeoniflorin inhibits the migration- and invasion-promoting capacities of GCAFs by targeting microRNA-149 and IL-6. Paeoniflorin is potentially a novel therapeutic agent against cancer microenvironment.

Astragaloside IV inhibits pathological functions of gastric cancer-associated fibroblasts.

AIM: To investigate the inhibitory effect of astragaloside IV on the pathological functions of cancer-associated fibroblasts, and to explore the underlying mechanism. METHODS: Paired gastric normal fibroblast (GNF) and gastric cancer-associated fibroblast (GCAF) cultures were established from resected tissues. GCAFs were treated with vehicle control or different concentrations of astragaloside IV. Conditioned media were prepared from GNFs, GCAFs, control-treated GCAFs, and astragaloside IV-treated GCAFs, and used to culture BGC-823 human gastric cancer cells. Proliferation, migration and invasion capacities of BGC-823 cells were determined by MTT, wound healing, and Transwell invasion assays, respectively. The action mechanism of astragaloside IV was investigated by detecting the expression of microRNAs and the expression and secretion of the oncogenic factor, macrophage colony-stimulating factor (M-CSF), and the tumor suppressive factor, tissue inhibitor of metalloproteinase 2 (TIMP2), in different groups of GCAFs. The expression of the oncogenic pluripotency factors SOX2 and NANOG in BGC-823 cells cultured with different conditioned media was also examined. RESULTS: GCAFs displayed higher capacities to induce BGC-823 cell proliferation, migration, and invasion than GNFs (P < 0.01). Astragaloside IV treatment strongly inhibited the proliferation-, migration- and invasion-promoting capacities of GCAFs (P < 0.05 for 10 µmol/L, P < 0.01 for 20 µmol/L and 40 µmol/L). Compared with GNFs, GCAFs expressed a lower level of microRNA-214 (P < 0.01) and a higher level of microRNA-301a (P < 0.01). Astragaloside IV treatment significantly up-regulated microRNA-214 expression (P < 0.01) and down-regulated microRNA-301a expression (P < 0.01) in GCAFs. Reestablishing the microRNA expression balance subsequently suppressed M-CSF production (P < 0.01) and secretion (P < 0.05), and elevated TIMP2 production (P < 0.01) and secretion (P < 0.05). Consequently, the ability of GCAFs to increase SOX2 and NANOG expression in BGC-823 cells was abolished by astragaloside IV. CONCLUSION: Astragaloside IV can inhibit the pathological functions of GCAFs by correcting their dysregulation of microRNA expression, and it is promisingly a potent therapeutic agent regulating tumor microenvironment.

[Effect of modified sijunzi decoction on the bone metabolism of adriamycin induced nephropathy rats].

OBJECTIVE: To explore the effect of Modified Sijunzi Decoction (MSD) on the bone metabolism of prednisone intervened adriamycin-induced nephropathy rats. METHODS: The adriamycin-induced nephropathy rat model was prepared. Totally 50 SD rats were randomly divide into five groups, i.e., the model group, the hormone group, the Chinese medicine (CM) group, the CM + hormone group, and the normal control group. The 24-h urine samples were collected on the 7th, 21st, and 35th day after modeling. The 24-h urine protein was measured by biuret colorimetry. Serum levels of osteoprotegerin (OPG), receptor activator of nuclear factor-kappaB ligand (RANKL), osteocalcin (BGP), and tartrate-resistant acid phosphatase (TRACP) were determined by ELISA. Expressions of OPG and RANKL in the tibia tissue were detected using real-time quantitative PCR and Western blot. RESULTS: (1) Compared with the normal control group, the 24-h urine protein increased in each group on the 7th, 21st, and 35th day (P < 0.05, P < 0.01). Compared with the model group, the 24-h urinary protein decreased in the hormone group and the CM + hormone group (P < 0.05, P < 0.01). The decrement was more obvious along with the treatment time went by (P < 0.05, P < 0.01). There was statistical difference in the reduction of urine protein on the 35th day between the CM group and the model group (P < 0.05). (2) Compared with the 21st-day of the same group, the serum levels of TRACP and RANKL increased (P < 0.05, P < 0.01). Compared with the model group, the serum levels of the TRACP and RANKL increased (P < 0.05, P < 0.01), OPG and BGP decreased (P < 0.05, P < 0.01) in the hormone group. Compared with the CM group at the same period, serum OPG level decreased and the RANKL level increased in the hormone group and the CM + hormone group (P < 0.05, P < 0.01). Besides, the serum level of TRACP increased and BGP decreased (P < 0.05, P < 0.01). Compared with the hormone group at the same period, OPG and BGP increased (P < 0.05, P < 0.01), RANKL decreased (P < 0.01) in the CM + hormone group. On the 35th day TRACP decreased (P < 0.01). (3) Compared with the normal group, mRNA expressions of OPG and RANKL on the 21st day increased (P < 0.05, P < 0.01), mRNA expressions of OPG and RANKL on the 35th day decreased in the model group (P < 0.01). Compared with the CM group at the same period, OPG mRNA expression decreased (P < 0.01) and RANKL mRNA expression increased in the hormone group (P < 0.05). OPG mRNA expression decreased in the CM +hormone group (P < 0.05). (4) Compared with the hormone group on the 21st day, the OPG level decreased and the RANKL protein increased (both P < 0.05). RANKL decreased in the CM + hormone group (P < 0.05). Compared with the model group at the same period, OPG decreased and RANKL increased in the hormone group (P < 0.01). Compared with the CM group at the same period, OPG decreased (P < 0.01), RANKL increased (P < 0.01) in the hormone group and the CM + hormone group. Compared with the hormone group at the same period, OPG increased and RANKL decreased in the CM + hormone group (both P < 0.01). CONCLUSIONS: Prednisone could induce osteoporosis through the OPG/RANKL/RANK pathway. MSZ could slow down the formation of prednisone-induced osteoporosis through promoting osteoblast differentiation, and inhibiting osteoclastogenesis.