Cellular APOBEC3A deaminase drives mutations in the SARS-CoV-2 genome.

The number of genetic variations in the SARS-CoV-2 genome has been increasing primarily due to continuous viral mutations. Here, we report that the human APOBEC3A (A3A) cytidine deaminase plays a critical role in the induction of C-to-U substitutions in the SARS-CoV-2 genome. Bioinformatic analysis of the chronological genetic changes in a sequence database indicated that the largest UC-to-UU mutation signature, consistent with APOBEC-recognized nucleotide motifs, was predominant in single-stranded RNA regions of the viral genome. In SARS-CoV-2-infected cells, exogenous expression of A3A but not expression of other APOBEC proteins induced UC-to-UU mutations in viral RNA (vRNA). Additionally, the mutated C bases were often located at the tips in bulge or loop regions in the vRNA secondary structure. Interestingly, A3A mRNA expression was drastically increased by interferons (IFNs) and tumour necrosis factor-α (TNF-α) in epithelial cells derived from the respiratory system, a site of efficient SARS-CoV-2 replication. Moreover, the UC-to-UU mutation rate was increased in SARS-CoV-2 produced from lung epithelial cells treated with IFN-ß and TNF-α, but not from CRISPR/Cas9-based A3A knockout cells. Collectively, these findings demonstrate that A3A is a primary host factor that drives mutations in the SARS-CoV-2 RNA genome via RNA editing.

Changes in graft function after living donor kidney transplantation in children.

BACKGROUND: This study aimed to evaluate the change in graft function in two groups stratified by the estimated glomerular filtration rate (eGFR) at 1 month after transplantation (eGFR-1 M) in pediatric living donor kidney transplant recipients. METHODS: Forty-three pediatric recipients were classified as those with an eGFR-1 M ≥ 90 mL/min/1.73 m2 (n = 19; high eGFR group) or those with an eGFR-1 M of 60-89 mL/min/1.73 m2 (n = 24; middle eGFR group). In the two groups, changes in the eGFR were retrospectively evaluated for 5 years after kidney transplantation. RESULTS: The mean recipient age at transplantation in the high/middle eGFR group was 6.1 ± 3.4/7.8 ± 4.0 years (P = 0.14). The mean eGFR-1, -12, and -60 M (mL/min/1.73 m2) in the high/middle eGFR group were 106.8 ± 2.99/78.5 ± 1.52 (P < 0.001), 79.3 ± 3.22/62.7 ± 2.38 (P < 0.001), and 73.1 ± 4.16/59.2 ± 2.79 (P = 0.006), respectively. The change in the mean eGFR remained mostly parallel in the two groups. In both groups, the eGFR significantly decreased only between 1 and 12 months after transplantation (P < 0.0001). Approximately 70% of the patients had an eGFR-60 M ≥ 60 mL/min/1.73 m2. CONCLUSIONS: The high and middle eGFR groups showed a rapid decline in the eGFR by 1 year after transplantation, but the change thereafter was gradual. In pediatric living donor kidney transplant recipients, the eGFR was relatively well maintained up to 5 years after transplantation. A higher resolution version of the Graphical abstract is available as Supplementary information.

Long-term social outcome after pediatric kidney transplantation: a single-center experience.

BACKGROUND: Patient and graft survival rates after pediatric kidney transplantation have improved recently. Therefore, the quality of life or social outcome after kidney transplantation has become important for patients and their families. METHODS: Patients who underwent kidney transplantation at < 18 years old and were observed for > 10 years were included in this study. The median age at first kidney transplantation was 9.2 (interquartile range [IQR] = 5.6-13.0) years; there were 56 males and 50 females. The median age at last follow-up was 29.9 (IQR = 22.2-36.0) years. We evaluated the patients' renal function, growth, professional status, and marital status at the last follow-up. RESULTS: The percentage of functioning grafts at the last follow-up was 81.1%; 73 patients (68.9%) had a first graft. The mean estimated GFR was 51.0 ± 20.5 mL/min/1.73 m2. Twenty patients received dialysis for graft failure. The mean final heights of the males and females were 158.1 ± 9.2 cm (- 2.2 standard deviations) and 149.1 ± 6.4 cm (- 1.7 standard deviations), respectively. Excluding 23 students, 63 patients (75.9%) were employed. Office worker was the most common profession. Twelve patients (14.5%) were unemployed. Of patients > 20 years old, 14 (16.7%), three males and 11 females, were married. Five females had one child each. CONCLUSIONS: The graft survival rate was favorable. The final height was short, particularly in male. The rate of employment was relatively high. The rate of marriage and having children were still low. Improving the social outcome is an important problem after pediatric kidney transplantation.

Specific mutations in the HIV-1 G-tract of the 3'-polypurine tract cause resistance to integrase strand transfer inhibitors.

BACKGROUND: In vitro selection experiments identified viruses resistant to integrase strand transfer inhibitors (INSTIs) carrying mutations in the G-tract (six guanosines) of the 3'-polypurine tract (3'-PPT). A clinical study also reported that mutations in the 3'-PPT were observed in a patient receiving dolutegravir monotherapy. However, recombinant viruses with the 3'-PPT mutations that were found in the clinical study were recently shown to be susceptible to INSTIs. OBJECTIVES: To identify the specific mutation(s) in the G-tract of the 3'-PPT for acquiring INSTI resistance, we constructed infectious clones bearing single or multiple mutations and systematically characterized the susceptibility of these clones to both first- and second-generation INSTIs. METHODS: The infectious clones were tested for their infectivity and susceptibility to INSTIs in a single-cycle assay using TZM-bl cells. RESULTS: A single mutation of thymidine (T) at the fifth position (GGG GTG) in the G-tract of the 3'-PPT had no effect on INSTI resistance. A double mutation, cytidine (C) or 'T' at the second position and 'T' at the fifth position (GCG GTG and GTG GTG), increased resistance to INSTIs, with the appearance of a plateau in the maximal percentage inhibition (MPI) of the dose-response curves, consistent with a non-competitive mechanism of inhibition. CONCLUSIONS: Mutations at the second and fifth positions in the G-tract of the 3'-PPT may result in complex resistance mechanism(s), rather than simply affecting INSTI binding at the IN active site.

Comparison of live donor predonation and post-transplant kidney volumes and glomerular size in pediatric patients weighing less than 15 kg - a retrospective study.

Renal transplantation of adult-size kidneys presents a size mismatch in small children. This study presents a comparison of live donor predonation and recipient post-transplant kidney volumes (k-vol) and glomerular size at 1 year after transplantation. We analyzed 47 pediatric renal transplant recipients weighing <15 kg between 2009 and 2017. The k-vol before and 1 year after transplantation and glomerular size at implant and 1 year post-transplant were evaluated. We estimated the relationships between these changes and graft function, and the factors associated with k-vol. Pretransplant k-vol was 158.1 ± 25.1 ml, and the k-vol at 1 year post-transplant was significantly reduced by -17.2% to 132.3 ± 27.3 ml (P < 0.001). Implant glomerular size showed the diameter was 165.3 ± 15.1 µm and the area 20 737.1 ± 3230.6 µm2 . One-year post-transplant, the glomerular diameter was 150.6 ± 11.4 µm and the area 17 428.3 ± 2577.9 µm2 , significantly reduced compared with implantation values (both P < 0.001). The change in k-vol was affected by pretransplant abdominal cavity (ml/200 ml cavity volume, partial regression coefficient = 0.029, SE = 0.009, P = 0.004) and recipient's weight gain (ml/5% of weight gain, partial regression coefficient = 0.020, SE = 0.006, P = 0.002). In small pediatric transplants, an adult-size kidney is acceptable with reduction in k-vol. Moreover, the post-transplant k-vol might be regulated by pretransplant physique and post-transplant somatic growth.

ABO-incompatible pediatric kidney transplantation without antibody removal.

BACKGROUND: Because of the severe shortage of suitable deceased donors, ABO-incompatible living donor kidney transplantation (ABOi LDKT) is performed even in pediatric recipients in Japan. We performed pediatric ABOi LDKT using rituximab without anti-A/B antibody removal. METHODS: Thirteen pediatric recipients (mean age 7.4, range 3.4-15.7, four females) whose baseline anti-A/B IgG titers were ≤ × 64 underwent ABOi LDKT without antibody removal and splenectomy between July 2013 and April 2017 at Toho University. Mycophenolate mofetil (MMF) was initiated on day - 10. Rituximab (100 mg) was administered twice. Basiliximab and triple maintenance immunosuppression (calcineurin inhibitor, MMF, and steroids) were administered. Protocol biopsy was performed at 3 months and 1 year after transplantation. We retrospectively compared the clinical outcomes between these recipients and 37 children (mean age 9.0, range 2.6-18.9, 15 female) who underwent ABO-compatible (ABOc) LDKT during the same period. RESULTS: The mean follow-up periods of ABOi and ABOc groups were 31.9 ± 13.5 and 28.8 ± 14.4 months, respectively. In the ABOi group, no clinical acute rejection (AR) was noted and subclinical AR was observed in four patients without evidence of acute antibody-mediated rejection. In the ABOc group, clinical and subclinical AR developed in 3 and 10 patients, respectively. No significant difference was identified for the mean eGFR between the ABOi and ABOc groups (98.3 ± 48.8 vs. 86.9 ± 39.4, P = 0.452 at 3 months; 78.2 ± 21.2 vs. 79.7 ± 21.3, at 1 year, P = 0.830). Death-censored graft survival at follow-up was 100% in the ABOi group and 94.6% in the ABOc group. Patient survival during the follow-up period in both the groups was 100%. Late-onset neutropenia (LON) requiring granulocyte colony-stimulating factor occurred more frequently in the ABOi group than in the ABOc group (4 vs. 0 patients) (P < 0.001). CONCLUSIONS: Pre- and post-transplantation antibody removal is not a prerequisite for successful pediatric ABOi LDKT, at least in patients with a low anti-A/B IgG antibody titer. However, LON caused by rituximab should be monitored.

Surgical Challenge in Pediatric Kidney Transplant Vascular Anastomosis.

During pediatric kidney transplant, surgical challenges occasionally occur. In particular, vascular anastomosis should be considered for children with small body weight < 12 kg, multiple renal arteries, vascular anomaly, and inferior vena cava occlusion. In pediatric patients, a living-donor renal graft is usually donated from a parent. Therefore, the renal artery and vein are too large to be anastomosed with the recipient's internal iliac artery and external iliac vein. In children who are > 12 kg, the renal artery and vein could be anastomosed with the external iliac artery and the external iliac vein. In children who are < 10 kg, the renal artery and vein should be anastomosed directly with the aorta and inferior vena cava. A pediatric transplant surgeon should consider arterial and venous anastomosis sites before transplant surgery. In small children with partial or total inferior vena cava occlusion, the venous anastomosis site should be evaluated. If the graft is placed on the left side, a venous graft must be used as a bridge between the renal vein and inferior vena cava. In 13 kidney transplants in children with inferior vena cava occlusion, 7 were on the left and 6 were on the right side. A patent segment of the inferior vena cava, the left original renal vein, an ascending lumbar vein, an azygos vein, the first graft renal vein, and a portal vein were used for venous anastomosis in 6, 2, 2, 1, 1 and 1 recipient, respectively. One child had graft loss due to renal vein thrombosis and one died of hemorrhage immediately posttransplant. Three had grafts with relatively long-term function, but these were lost due to chronic allograft nephropathy 100, 122, and 137 months posttransplant. However, the other 8 recipients have so far maintained graft function from 6 to 138 months since transplant.

Surgical Challenge in Pediatric Kidney Transplant: Lower Urinary Tract Abnormality.

Lower urinary tract abnormalities are difficult to resolve in pediatric kidney transplant patients. Measure of residual urine, voiding cystourethrography, retrograde urethrography, cystometry, electromyography of urethral external sphincter muscle, urethrometry, and uroflowmetry are the primary methods for evaluation of lower urinary tract abnormalities. Endoscopic resection or ablation of urethral valves is required in children with posterior urethral valve to treat obstruction, but bladder function does not always recover and may deteriorate to end-stage renal failure even after the obstruction is released. This bladder dysfunction in posterior urethral valve defines valve bladder syndrome. Vesicoureteral reflux caused by high vesical pressure can cause even worse renal graft function posttransplant. In our patient group, urinary diversion occurred with Mitrofanoff conduit using an appendix in 6 children, a Yang-Monti channel conduit using ileum in 1 patient, with cystostomy in 3 children, and with augmented cystoplasty in 9 children before or simultaneously with kidney transplant. These procedures should be selected based on the type of lower urinary tract abnormality including bladder function. Recently, we have preferred a continent diversion for self-catheterization in children with lower urinary tract abnormalities. We have conducted 9 augmented cystoplasty procedures using a portion of the sigmoid colon or ileum. Seventeen children retained their own bladders when the transplant ureter was implanted. Most patients needed clean intermittent catheterization, depending on the residual urine volume and a bladder function. Ten-year graft survival rate in kidney transplant in our department is 98% in 36 children with lower urinary tract abnormalities. Lower urinary tract abnormality is not always a risk factor for pediatric kidney transplant; however, a preoperative evaluation is important to choose the best option for urinary diversion.

Increase of serum vascular endothelial growth factors in wet beriberi: two case reports.

Beriberi is a disease caused by thiamine deficiency resulting in peripheral neuropathy and myocardial dysfunction. Increases in vascular endothelial growth factor (VEGF) are seen in polyneuropathy, organomegaly, endocrinopathy, monoclonal gammopathy and skin changes, called POEMS syndrome. We present herein two cases of wet beriberi accompanied by a moderate increase in VEGF level. Serum VEGF decreased after treatment in both cases. Our experience with these cases suggests that beriberi should be considered in the differential diagnosis of polyneuropathy with a moderate increase in serum VEGF, and that the serum VEGF level may be a therapeutic marker for beriberi.