Cytokinetic engineering enhances the secretory production of recombinant human lysozyme in Komagataella phaffii.

BACKGROUND: Human lysozyme (hLYZ) is a natural antibacterial protein with broad applications in food and pharmaceutical industries. Recombinant production of hLYZ in Komagataella phaffii (K. phaffii) has attracted considerable attention, but there are very limited strategies for its hyper-production in yeast. RESULTS: Here through Atmospheric and Room Temperature Plasma (ARTP)-based mutagenesis and transcriptomic analysis, the expression of two genes MYO1 and IQG1 encoding the cytokinesis core proteins was identified downregulated along with higher hLYZ production. Deletion of either gene caused severe cytokinesis defects, but significantly enhanced hLYZ production. The highest hLYZ yield of 1,052,444 ± 23,667 U/mL bioactivity and 4.12 ± 0.11 g/L total protein concentration were obtained after high-density fed-batch fermentation in the Δmyo1 mutant, representing the best production of hLYZ in yeast. Furthermore, O-linked mannose glycans were characterized on this recombinant hLYZ. CONCLUSIONS: Our work suggests that cytokinesis-based morphology engineering is an effective way to enhance the production of hLYZ in K. phaffii.

Immunoglobulin a vasculitis with testicular/epididymal involvement in children: A retrospective study of a ten-year period.

The clinical characteristics and risk factors for testicular/epididymal involvement in 73 children with immunoglobulin A vasculitis (IgAV) who were admitted to our hospital between January 2012 and November 2022 were reviewed. The demographic data, laboratory parameters, and follow-up data of the patients were compared to those of 146 males without testicular/epididymal involvement. A logistic regression analysis was performed to determine the variables associated with testicular/epididymal involvement. The prevalence of testicular/epididymal involvement among male patients with IgAV was 1.3% (73/5,556). Increased blood flow in the testes and/or epididymis on ultrasound was found in 71 patients. The remaining two patients underwent surgical exploration for loss or reduction of testicular blood flow. One patient underwent orchiectomy for intraoperative confirmation of complete right testicular infarction. Pathological findings revealed IgA immune complex deposition in the testis. Patient age (odds ratio [OR] = 0.792; 95% confidence interval [CI]: 0.682-0.919, p = 0.002), platelet count (OR = 1.011; 95% CI: 1.002-1.020, p = 0.013), and immunoglobulin M (IgM) levels (OR = 0.236; 95% CI: 0.091-0.608, p = 0.003) were strongly associated with the occurrence of testicular/epididymal involvement in IgAV. Therefore, young age, increased platelet count, and low IgM levels in patients with IgAV are potential risk factors for testicular/epididymal involvement. Doppler ultrasound can help differentiate IgAV from acute scrotum. Most patients with testicular/epididymal involvement have good prognoses, although serious complications such as testicular infarction may occur.

[Molecular mechanism underlying difference of astragaloside â £ content in imitating wild and cultivated Astragalus mongholicus].

The difference of astragaloside Ⅳ content and the expression of its biosynthesis related genes in imitating wild Astragalus mongolicus(IWA) and cultivated A.mongolicus(CA) under different growth years were systematically compared and analyzed.Then the key enzyme genes affected the difference of astragaloside Ⅳ content in the above two A.mongolicus were screened.High-perfo-rmance liquid chromatography(HPLC)was used to determine the content of astragaloside Ⅳ in A.mongolicusunderthe above two diffe-rent growth patterns.Based on the Illumina HiSeq and PacBio high-throughput sequencing platforms, thesecond-and third-generation transcriptome sequencing(RNA-Seq)databaseof the two A.mongolicuswas constructed.The related enzyme genes in the biosynthetic pathway of astragaloside Ⅳ were screened and verified byquantitative reverse transcriptase polymerase chain reaction(RT-qPCR).The RNA-sequencing(RNA-Seq) and RT-qPCR data of each gene were subjected to correlation analysis and trend analysis.The results showed that the variation trend of astragaloside Ⅳ contentby HPLC wasthe same as that of genes by RNA-Seq and RT-qPCR in 1-4 year IWA and 1-2 year CA.The trend level of astragaloside Ⅳ contentwas lower in 2-year IWA than 1-year IWA.Compared with 2-year IWA, 3-year IWA had an upward trend, while 4-year IWA hada downward trend versus 3-year IWA.Additionally, 1-year CA had increased trendthan 2-year CA.However, the content of astragaloside Ⅳ in 5-year IWA was higher than that of 6-year IWA, which wasinconsistent with the findings of RNA-Seq and RT-qPCR.This study preliminarily clarifiedthat the difference of astragaloside Ⅳ contentin 1-4 year IWA and 1-2 year CA wasclosely related to the expression of the upstream and midstream genes(MVK, CMK, PMK, MVD, SS) in the biosynthetic pathway.The results facilitate the production and planting of Radix Astragali seu Hedysari.

[Unexpected cutaneous purpura in an infant].

A two-month-old boy visited the hospital due to unexpected cutaneous purpura and thrombocytopenia for 2 days. The physical examination revealed a purple mass on the back. The soft tissue color Doppler ultrasound showed rich blood signals in the tissue, and the results of bone marrow puncture indicated an increased number of megakaryocytes. After the treatment with hormone and gamma globulin, the platelet count rapidly increased and maintained at a normal level. Meanwhile, the boy was given oral administration of propranolol. He was followed up for 4 months and the volume of the mass on the back was reduced significantly. He had a definite diagnosis of hemangioma and immune thrombocytopenia. As for the patients with hemangioma complicated by thrombocytopenia, knowledge of Kasabach-Merritt syndrome should be enhanced and there should be a clarification of the association between thrombocytopenia and hemangioma. There should also be an alertness for thrombocytopenia of other causes.

[Misdiagnosis of pancoast cancer: analysis of 26 cases].

OBJECTIVE: To further understand the clinical features and the causes of misdiagnosis of pancoast cancer. METHODS: The data of 26 cases of pancoast cancer from 1999 - 2006 were collected for a retrospective review. The clinical and imaging features, and the causes underlying misdiagnosis were analyzed. RESULTS: There were 24 males and 2 females (mean age 59 ± 6, range 44 - 70 years). The earliest clinical features of pancoast cancer included painful shoulder and back in 25 cases, upper extremity pain, palsy and numbness in 18 cases. Early respiratory symptoms such as cough and sputum were recorded in 5 cases, Horner syndrome in 2 cases, and hoarseness in 1 case. Misdiagnosis was made in 19 of the 26 cases: 13 as periarthritis humeroscapularis, 3 as cervical osteoarthritis and another 3 as pulmonary tuberculosis. CT scanning showed soft tissue masses at the apical segments of the lungs in all the cases (26/26). Three cases were misdiagnosed as "pulmonary tuberculosis" according to the images of X-ray, and another 3 went unnoticed. CONCLUSIONS: Painful shoulder and back were the most common symptoms of pancoast cancer. pancoast cancer was often diagnosed at an advanced stage, and misdiagnosis was common. Thorax CT was the most valuable method for the detection of Pancoast cancer.