Peripheral blood cell telomere length measurements indicate that hematopoietic stem cell turnover is not significantly increased in whole blood and apheresis PLT donors

BACKGROUND: The telomere length (TEL) of peripheral blood leukocytes (PBLs) can be used to estimate hematopoietic stem cell turnover. The current study investigated whether the repetitive stimulation of the hematopoietic system caused by regular whole blood (WB) and PLT donations would affect PBL TEL. STUDY DESIGN AND METHODS: PBLs were obtained from healthy donors (n=94) with a history of at least 3 years of WB donation (median, 7.7 years; range, 3.0-43.0 years) plus additional apheresis PLT donations. The median (range) numbers of WB and PLT donations were 22.0 (6.0-194.0) and 42.0 (7.0-336.0), respectively. Additionally, samples were obtained from healthy nondonors (n=47). PBL TEL was measured with fluorescence in situ hybridization and flow cytometry (flow-FISH). Flow-FISH results were expressed in molecular equivalents of soluble fluorochrome units (MESF; 1000 MESF=1 kMESF) either as absolute (TEL) or as age-adjusted TEL (DeltaTEL). RESULTS: Donor granulocyte and lymphocyte TELs were 12.6 +/- 0.3 (mean +/- SEM) and 13.2 +/- 0.3 kMESF, respectively. No differences were observed when compared with corresponding nondonor data (granulocytes, 12.5 +/- 0.4 kMESF; lymphocytes, 13.6 +/- 0.5 kMESF). Furthermore, DeltaTEL values did not differ between the two groups and were not different from previously established reference values. In addition, neither donor data for age-adjusted TEL for granulocytes nor DeltaTEL for lymphocytes were correlated with either total years or total numbers of WB and/or PLT donations. CONCLUSION: Long-term WB and PLT donation does not affect PBW TEL as measured by flow-FISH, arguing against a significantly increased stem cell turnover.     

Eradication of severe neonatal systemic candidiasis with amphotericin B lipid complex

OBJECTIVE: To report the successful use of amphotericin B lipid complex in treating severe systemic candidiasis in a very-low-birth-weight infant. CASE SUMMARY: A preterm female infant, born at 25 weeks' gestational age with a birth weight of 870 g, had received full supportive care in the neonatal intensive care unit (NICU), including mechanical ventilation, total parenteral nutrition, and placement of central venous catheters. At seven weeks of age, she developed severe disseminated candidiasis, which failed to respond to conventional amphotericin B and fluconazole therapy. Her progressive deterioration was reversed only after amphotericin B lipid complex (A-complex) was substituted for conventional amphotericin B. The improvement in her condition was impressive, and she made a full recovery without any adverse effect. DISCUSSION: With increased reliance on invasive technologies for life support, systemic candida infections have become increasingly common among premature infants in the NICU. Such infections are potentially fatal for the high-risk neonate. A literature review shows limited documentation of the use of lipid-based formulations of amphotericin B, especially A-complex, in preterm infants. However, the collective experience with these products appears to show that they are effective and cause fewer adverse effects than conventional amphotericin B. The infant reported here had shown progressive deterioration from disseminated candidiasis until conventional amphotericin B therapy was replaced with A-complex. Her recovery corresponded to the clearance of the candidemia. CONCLUSIONS: With favorable results and increasing experience with lipid-based formulations of amphotericin B, it is reasonable to consider these new formulations as therapy for candidemia in preterm infants who are at a high risk of nephrotoxicity or who have failed conventional therapy.     

Endoscopic ultrasonographic study of the azygos vein before and after endoscopic obliteration of esophagogastric varices by injection sclerotherapy

BACKGROUND AND STUDY AIMS: The azygos vein plays an important role as a drainage system for the superior portosystemic collateral circulation in portal hypertensive patients. Endoscopic ultrasonography (EUS) and Doppler EUS allow the performance of hemodynamic studies of the azygos vein. In this study, we observed the changes in the azygos vein which occur with variceal obliteration by endoscopic injection sclerotherapy (EIS). PATIENTS AND METHODS: We recruited patients with portal hypertension and bleeding varices who were not on portal pressure-lowering agents and who were scheduled for the EIS program. EUS was performed in these patients to study the azygos vein at the start of EIS. The azygos vein diameter, maximal velocity (Vmax), and blood flow volume index (BFVI) were measured. After variceal obliteration and within 1 week, another EUS study of the azygos vein was carried out. RESULTS: Out of 40 patients recruited into the study variceal obliteration and EUS assessment of the azygos vein, within 1 week of obliteration, was achieved in 33. We noticed a significant increase in azygos vein diameter (P<0.001) and BFVI (P=0.001) following variceal obliteration. No significant change was observed in Vmax (P>0.05). In one patient, marked caliber irregularities were observed in the azygos vein after variceal obliteration. CONCLUSIONS: Using EUS and Doppler EUS, hemodynamic studies of the azygos vein blood flow can be performed, allowing the monitoring of the effects of EIS and variceal obliteration on the superior portosystemic collateral circulation. The clinical significance of the observed changes in azygos blood flow that occur with variceal obliteration should be investigated in further studies and correlated with short-term and long-term outcome.     

Donor-derived alloantibodies and passenger lymphocyte syndrome in two of four patients who received different organs from the same donor

BACKGROUND: Reported here is the occurrence of RBC alloimmunization in two of four patients who received different organs from an immunized donor. STUDY DESIGN AND METHODS: The donor, a 58-year-old woman, was group O D+, K-, and Fy(a-). Initially, her serum contained only a K antibody. After blood transfusion, a second antibody (anti-Fy(a)) could also be identified. The liver was given to a group O D+, K-, Fy(a+) patient; the pancreas and one kidney to a group O D+, K-, Fy(a+) patient; the heart to a group A D+, K-, Fy(a-) patient; and the other kidney to a group B D+, K-, Fy(a+) patient. RBC grouping and antibody screening were performed by standard techniques. Lymphoid microchimerism in the peripheral blood of the recipients was analyzed by flow cytometry and nested PCR. RESULTS: None of the recipients had irregular RBC alloantibodies at the time of transplantation. After the transplant, anti-K became detectable in the serum of the liver recipient, and anti-Fy(a) could be eluted from the RBCs of the liver recipient and the pancreas-kidney recipient. The latter patient also developed mild hemolysis, and his Hb dropped to 8 g per dL on posttransplant Day 9. Donor-derived lymphocytes were detectable by flow cytometry in the peripheral blood of the liver recipient and the pancreas-kidney recipient until Days 8 and 63, respectively, whereas no lymphoid chimerism could be demonstrated in the heart recipient. PCR chimerism analyses were positive in all three recipients over the whole observation period of 97 postoperative days. CONCLUSION: The amount of cotransplanted lymphoid tissue may correlate with the extent of peripheral lymphoid microchimerism and the antibody-formation capacity in solid organ transplantation.     

Magnetised Thermo Responsive Lipid Vehicles for Targeted and Controlled Lung Drug Delivery

Purpose

Conditions such as lung cancer currently lack non-invasively targetable and controlled release topical inhalational therapies. Superparamagnetic iron-oxide nanoparticles (SPIONs) have shown promising results as a targetable therapy. We aimed to fabricate and test the in-vitro performance of particles with SPION and drug within a lipid matrix as a potentially targetable and thermo-sensitive inhalable drug-delivery system.

Methods

Budesonide and SPIONs were incorporated into lipid particles using oil-in-water emulsification. Particles size, chemical composition, responsiveness to magnetic field, thermosensitiveness and inhalation performance in-vitro were investigated.

Results

Particles of average diameter 2?C4???m with budesonide and SPIONs inside the lipid matrix responded to a magnetic field with 100% extraction at a distance of 5?mm. Formulations were shown to have accelerated rate of drug release at hyperthermic temperatures (45°C)??controlled release. The produced inhalation dry powder presented promising inhalation performance, with an inhalable fine particle fraction of 30%.

Conclusions

The lipid system presented thermo-sensitive characteristics, suitable for controlled delivery, the model drug and SPION loaded lipid system was magnetically active and movable using simple permanent magnets, and the system demonstrates promise as an effective drug vehicle in targeted and controlled inhalation therapy.     

Amino acid chloramine damage to proliferating cell nuclear antigen in mammalian cells

Amino acid chloramines (AACLs) are reactive secondary products of activated neutrophils. To understand AACL damage in cell nuclei, we exploited proliferating cell nuclear antigen (PCNA) as a nuclear protein damage reporter, using western blotting and mass spectrometry. Chloramines of proline, arginine, and glycine caused significant damage to PCNA in cells. Chloramines of taurine and histidine caused slight damage to PCNA in cells. Other AACLs caused no PCNA damage in intact cells. Evidence supports a sulfonamide, sulfinamide, or sulfenamide crosslinking mechanism involving cysteine 148 at the PCNA subunit interface, methionine sulfoxide formation as the basis of electrophoretic mobility shifting, and tyrosine and/or methionine residues as the likely targets of AACL damage to the PCNA antibody epitope. An interstitial fluid model experiment showed that physiological amino acids can mediate HOCl damage to PCNA in the presence of proteins that would otherwise completely quench the HOCl. CONCLUSION: PCNA is a sensitive biomarker of AACL damage in cell nuclei. Arginine chloramine and proline chloramine, or reactive species derived from them, were shown to enter cells and damage PCNA. Amino acids were shown to have at least two different mechanisms for suppressing PCNA damage in cells by their corresponding AACLs. Cysteine 148 was shown to be essential for PCNA subunit crosslinking by AACLs, and a crosslinking mechanism was proposed.     

An outbreak of CTX-M-15-producing Klebsiella pneumoniae isolates in an intensive care unit of a teaching hospital in Kuwait

Objective: This study reports an outbreak of Klebsiella pneumoniae infections in 14 patients during a 2-month period (August–September, 2008) in the intensive care unit (ICU) of a teaching hospital in Kuwait. Materials and Methods: The clinical sources were blood (9), urine (3) and respiratory secretions (2) identified by the automated VITEK-2 ID System. Susceptibility testing was performed by the E-test method. Extended-spectrum β-lactamase (ESBL) production was assessed using the ESBL E-test and confirmed by PCR. Carriage of bla genes was determined by PCR and sequence analysis. The transferability of resistance phenotypes was demonstrated by conjugation experiments and clonal relatedness was determined by PFGE. Results: The isolates were susceptible to imipenem, meropenem, and tigecycline and produced ESBL. All isolates yielded an amplicon of 499 bp with universal consensus primers (MA primers). DNA sequence analysis showed that they all harboured bla_CTX-M-15and bla_TEM-1genes. The environmental isolate obtained from a suction machine was also CTX-M-15/TEM-1 producer. The resistance phenotypes were transferrable to the Escherichia coli J53^r strain. PFGE, revealed two clones, A and B, related with a Dice coefficient of >94.1%. A mortality rate of 21.4% was recorded. Conclusion: The outbreak was contained by robust and aggressive infection control measures. This study highlights the first outbreak of CTX-M-15-producing K. pneumoniae associated with high mortality in an adult medical ICU in Kuwait.     

Low expression of Granzyme B in colorectal cancer is associated with signs of early metastastic invasion

Salama P, Phillips M, Platell C & Iacopetta B
(2011) Histopathology 59 , 207–215 Low expression of Granzyme B in colorectal cancer is associated with signs of early metastastic invasion Aims: Tumour‐infiltrating forkhead box P3 (FoxP3⁺) regulatory T cells (T_regs) have stronger prognostic significance than cytotoxic CD8⁺ T cells in colorectal cancer (CRC). Because there is evidence that some tumour‐infiltrating CD8⁺ T cells may be inactive, the present study aimed to investigate the prognostic significance of Granzyme B, one of the major effector molecules of T cells. Methods and results: A tissue microarray containing 963 CRCs was stained immunohistochemically for Granzyme B and the level of expression quantified by digital image analysis. Granzyme B expression was higher in tumours with microsatellite instability (P < 0.0001), a dense lymphocytic infiltrate (P < 0.0001) and location in the proximal colon (P = 0.009), but lower in tumours with vascular invasion (P = 0.007), perineural invasion (P =0.041) and positive nodal status (P < 0.001). Elevated expression of Granzyme B was associated with improved survival on univariate analysis (hazard ratio = 0.65; 95% confidence interval 0.51–0.84; P = 0.001), but not in a multivariate model that included stage, vascular invasion and FoxP3⁺ T_reg cell density. Conclusions: Low expression of Granzyme B was associated with early signs of metastasis in CRC. The stronger prognostic significance of FoxP3⁺ T_regs is in keeping with animal models that suggest these cells act as gatekeepers for the release of Granzyme B from CD8⁺ T cells.     

Identification of a novel HLA-C*02 allele, HLA-C*02:02:09

We report HLA-C*02:02:09 as a novel allele with a transition C->T at position 354.