Streamlined human antibody generation and optimization by exploiting designed immunoglobulin loci in a B cell line

Monoclonal antibodies (mAbs) are widely utilized as therapeutic drugs for various diseases, such as cancer, autoimmune diseases, and infectious diseases. Using the avian-derived B cell line DT40, we previously developed an antibody display technology, namely, the ADLib system, which rapidly generates antigen-specific mAbs. Here, we report the development of a human version of the ADLib system and showcase the streamlined generation and optimization of functional human mAbs. Tailored libraries were first constructed by replacing endogenous immunoglobulin genes with designed human counterparts. From these libraries, clones producing full-length human IgGs against distinct antigens can be isolated, as exemplified by the selection of antagonistic mAbs. Taking advantage of avian biology, effective affinity maturation was achieved in a straightforward manner by seamless diversification of the parental clones into secondary libraries followed by single-cell sorting, quickly affording mAbs with improved affinities and functionalities. Collectively, we demonstrate that the human ADLib system could serve as an integrative platform with unique diversity for rapid de novo generation and optimization of therapeutic or diagnostic antibody leads. Furthermore, our results suggest that libraries can be constructed by introducing exogenous genes into DT40 cells, indicating that the ADLib system has the potential to be applied for the rapid and effective directed evolution and optimization of proteins in various fields beyond biomedicine.     

Evaluation of the relationship between linezolid exposure and hyponatremia

IntroductionAims of this study were (a) to assess the development ratio of hyponatremia during treatment with linezolid and (b) to evaluate the relationship between the risk of hyponatremia and linezolid exposure and patient background.MethodClinical data including linezolid serum concentrations and serum sodium values were collected at Toyama University Hospital and Kyorin University Hospital. Data from 89 patients were used for the analysis, and a nadir serum sodium level ≤130 mmol/L during the treatment with linezolid was defined as hyponatremia. Mann-Whitney's U test was used to evaluate the effects of the area under the time-concentration curve (AUC) of linezolid at the nadir sodium level, clinical characteristics (e.g. laboratory data), and baseline serum sodium levels on the development of hyponatremia.ResultsThe hyponatremia was occurred in 21 of 89 patients (23.6%). Data are compared for baseline and nadir serum sodium levels of patients with and without hyponatremia. In both groups, nadir serum sodium levels were significantly different from those of the baseline values (P < 0.05). The values of AUC_0-12, accumulated AUC, baseline serum sodium levels and age were significantly different between patients with and without hyponatremia (P < 0.05).ConclusionsLinezolid exposure, age, and baseline sodium levels were detected as the risk factors for linezolid-related hyponatremia. Our findings suggest that regular monitoring of serum sodium levels is desirable during treatment with linezolid, especially for the elderly and patients with low serum sodium levels before the start of linezolid administration.     

Width and depth of resection for small colorectal polyps: hot versus cold snare polypectomy

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The effects of postural changes of baroreflex gain in normal and hypertensive pregnancies

In order to understand the changes of baroreflex gain due to postural changes in normal pregnancies, we measured percentage changes (% changes) in blood pressure (SBP, DBP), heart rate (HR), stroke volume (SV), cardiac output (CO), total peripheral resistance (TPR) as well as cardiac autonomic nervous function (HF as an index of parasympathetic and LF/HF as an index of sympathetic function) and compared these parameters in normal pregnancies with those found in hypertensive pregnancies, such as chronic hypertensive (CHP) and severe preeclamptic pregnancies (PE), in late pregnancy (after 32 wks). When the position was changed from supine to standing in normal and non-pregnant women, the % changes of HR, DBP, TPR and LF/HF were increased and SBP, SV, CO and HF were decreased. The % changes of these parameters, however, were gradually decreased as pregnancy progressed, especially after 20-24 wks of gestation. In hypertensive pregnancies, however, even in late pregnancy, the decreased SBP and increased TPR was still observed and the profound decrease of CO and SV and increase of TPR were characteristic in PE when compared to CHP.     

The effect of celiac plexus block on heart rate variability

Background

Celiac plexus block (CPB) can be used for treating intra-abdominal visceral pain syndromes. The celiac plexus is the largest plexus of the sympathetic nervous system. Several nerve blocks have a marked effect on autonomic nervous activity. Furthermore, stellate ganglion block changes cardiac autonomic nervous activity. Thus, CPB could influence the sympathetic activity of the cardiac plexus. The aim of the present study was to see whether CPB modulated heart rate variability (HRV) in patients with pancreatic cancer.

Methods

Twelve patients received neurolytic CPB using 14 ml absolute alcohol. Data recorded in a palm-sized electrocardiographic unit were analyzed for HRV.

Results

CPB using a neurolytic solution did not induce any significant changes in the low-frequency (LF)/high-frequency (HF) ratio of HRV (LF/HF, P = 0.4642). Furthermore, the procedure did not induce any significant changes in blood pressure (systolic, P = 0.5051; diastolic, P = 0.5180).

Conclusion

CPB did not induce any significant changes in HRV or hemodynamics.     

Simple and rapid detection of the porcine reproductive and respiratory syndrome virus from pig whole blood using filter paper

The combination of Flinders Technology Associates filter papers (FTA cards) and real-time PCR was examined to establish a simple and rapid technique for the detection of porcine reproductive and respiratory syndrome virus (PRRSV) from whole pig blood. A modified live PRRS vaccine was diluted with either sterilised saline or pig whole blood, and the suspensions were applied onto the FTA cards. The real-time RT-PCR detection of PRRSV was performed directly with the samples applied to the FTA card without the RNA extraction step. Six whole blood samples from at random selected piglets in the PRRSV infected farm were also assayed in this study. The expected PCR product was successfully amplified from either saline diluted or pig whole blood diluted vaccine. The same PCR ampliocon was detected from all blood samples assayed in this study. This study suggested that the combination of an FTA card and real-time PCR is a rapid and easy technique for the detection of PRRSV. This technique can remarkably shorten the time required for PRRSV detection from whole blood and makes the procedure much easier.