A respiratory monitoring device based on clavicular motion.

Respiratory rate is one of the key vital signs yet unlike temperature, heart rate or blood pressure, there is no simple and low cost measurement device for medical use. Here we discuss the development of a respiratory sensor based upon clavicular motion and the findings of a pilot study comparing respiratory rate readings derived from clavicular and thoracic motion with an expiratory breath flow reference sensor. Simultaneously sampled data from resting volunteers (n = 8) was analysed to determine the location of individual breaths in the data set and from these, breath periods and frequency were calculated. Clavicular sensor waveforms were found to be more consistent and of greater amplitude than those from the thoracic device, demonstrating good alignment with the reference waveform. On comparing breath by breath periods a close agreement was observed with the reference, with mean clavicular respiratory rate R(2) values of 0.89 (lateral) and 0.98 (longitudinal-axis). This pilot study demonstrates the viability of clavicular respiratory sensing. The sensor is unobtrusive, unaffected by bioelectrical or electrode problems and easier to determine and more consistent than thoracic motion sensing. With relatively basic signal conditioning and processing requirements, it could provide an ideal platform for a low-cost respiratory monitor.

Measuring the TREC ratio in dried blood spot samples: intra- and inter-filter paper cards reproducibility.

The level of T-cell receptor excision circles (TREC), which decline with advancing age in normal individuals, has recently gained interest as a reference marker for studies on premature or early immunosenescence under particular health conditions. In order to facilitate translational studies at population and clinical levels, essential for the understanding of how changes in TREC levels are associated with responsiveness of the immune system, we have developed and optimized a real-time polymerase chain reaction (qPCR) assay which quantifies the TREC ratio from dried blood spot (DBS) samples. The present study considers a fully automated procedure to purify DNA and amplify sequences of interests by means of qPCR from DBS samples collected in healthy adults. Both TREC:PBMC (peripheral blood mononuclear cell) and TREC:T-cell ratios were compared for intra- and inter individual reproducibility. Furthermore, the impact of the length of storage on the quality of the DNA generated was also analyzed. In conclusion we describe a fully automated procedure for extracting DNA and qPCR set up, which offers a high-precision, robust qPCR assay for the quantification of both TREC:T-cell ratio and TREC:PBMC from DBS samples.

How important is vitamin D in preventing infections?

Interaction with the immune system is one of the most recently established nonclassic effects of vitamin D (VitD). For many years, this was considered to be limited to granulomatous diseases in which synthesis of active 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) or calcitriol is known to be increased. However, recent reports have supported a role for 1,25(OH)2D3 in promoting normal function of the innate and adaptive immune systems. Crucially, these effects seem to be mediated not only by the endocrine function of circulating calcitriol but also via paracrine (i.e., refers to effects to adjacent or nearby cells) and/or intracrine activity (i.e., refers to a hormone acting inside a cell) of 1,25(OH)2D3 from its precursor 25(OH)D3, the main circulating metabolite of VitD. The ability of this vitamin to influence human immune responsiveness seems to be highly dependent on the 25(OH)D3 status of individuals and may lead to aberrant response to infection or even to autoimmunity in those who are lacking VitD. The potential health significance of this has been underlined by increasing awareness of impaired status in populations across the globe. This review will examine the current understanding of how VitD status may modulate the responsiveness of the human immune system. Furthermore, we discuss how it may play a role in host resistance to common pathogens and how effective is its supplementation for treatment or prevention of infectious diseases in humans.

Real time-PCR assay estimating the naive T-cell pool in whole blood and dried blood spot samples: pilot study in young adults.

Because of their central role orchestrating the immune response, the decrease in repertoire number and diversity of naïve T-cells is a significant feature of immnosenescence. Reflecting the effective naive T-cell pool, quantifying the sj-TREC ratio (number of signal joint T-cell receptor excision circles/10(5) T-cells) in blood samples suffers however from constraints. The most limiting one is the absolute requirement of the flow cytometry analysis of peripheral blood samples for the T-cell numeration. In order to make this ratio more accessible for clinical and epidemiological studies addressing how changes in responsiveness of the immune system lead to an increased susceptibility to various diseases and poorer response to vaccination, we have developed a rapid and simple method for the quantification of the sj-TREC ratio in whole blood and in dried blood spot (DBS) samples. This novel method is a QPCR analysis using fluorescently labelled sequence-specific probes both for quantifying sj-TREC and T-cell count and therefore eliminating the absolute necessity of the flow cytometer analysis. In this pilot study, we have compared the sj-TREC ratio we obtained with this novel method in whole blood and in DBS samples of 10 healthy volunteers with those obtained with the technique of reference and found that they are comparable.

Tracing thymic output in older individuals.

As a result of age-associated thymic atrophy, T cell production declines with age. Some studies suggest that production undergoes an exponential decline starting at birth, while others consider the decline to be in a biphasic manner with a rapid reduction in output occurring before middle age followed by a phase in which output declines at a regular, albeit much slower, rate. Both approaches provide estimations of the time of termination of thymic output, but on the basis of limited amounts of data. We have analysed blood from more than 200 individuals between the ages of 58 and 104 years to determine changes in thymic output using signal-joint T cell receptor excision circles (sjTREC)/T cells as our measure. To reduce any potential geographical or nutritional bias we have obtained samples from five different European countries. Our results reveal that while the absolute number of T cells per microlitre of blood does not change significantly across the age range we tested, the values of sjTREC per microlitre show wide variation and reveal an age-associated decline in thymic output. In addition we show gender differences, with notably higher thymic output in females than males at each decade. More importantly, we noted a significant decline in sjTREC/T cell levels in those more than 90 years of age in both males and females. Our results provide information about the potential end-point for thymic output and suggest that sjTREC analysis may be a biomarker of effective ageing.

Immunity in the elderly: the role of the thymus.

Adjustments to lifestyle including social and medical changes have led to human populations having increased longevity in many countries, producing shifts in the population demographics. Approximately half of the increase in the world's population by 2050 may be accounted for by the prolonged survival of those over the age of 60. It is possible to age in relatively good health, but this is rare and for the majority of individuals, growing old is associated with functional impairment, an increased risk of developing a degenerative condition, an increased susceptibility to disease and an increased risk of death. The ageing human population is one of the most urgent challenges facing us today. Changes in the immune system are considered to have a critical role in the decline seen with age, since many infectious diseases may no longer kill an individual, but may contribute to more subtle overall changes. So the impact of infections in older individuals should not be measured only in terms of direct mortality rates, but also by their contribution to the 'indirect' mortality rate and to changes in the quality of life. Taking a pragmatic approach, we need to understand the drivers for immune decline if we are to consider intervening therapeutically in this process. One of the central drivers to this process is age-linked atrophy of the thymus and reversal of this process may have a considerable role in reversing immune decline.

The management of side-effects during therapy for hepatitis C.

The results of interferon and ribavirin combination therapy for chronic hepatitis C infection have substantially improved in recent years, such that the majority of patients in randomized-controlled trials now achieve a sustained virological response. However, adverse effects are commonplace, often disabling and may lead to interruption or cessation of therapy with subsequent loss of efficacy. Constitutional, neuropsychiatric and haematological reactions have proved particularly troublesome. In this review, we discuss these adverse effects in more detail and highlight recent advances in their management.

Evidence of thymic reconstitution after highly active antiretroviral therapy in HIV-1 infection.

OBJECTIVES: We aimed to provide evidence of thymic reconstitution after highly active antiretroviral therapy (HAART) in HIV-1 infected patients and to correlate this with the restoration of peripheral naïve T cells. METHODS: Positron emission tomography (PET) enables definitive evidence of thymic activity, indicating functional potential. In this case study, a single patient who initiated HAART demonstrated reconstitution of the naïve T-cell pool and underwent thymic PET scans at baseline and 2 and 6 months following initiation of therapy. Two patients who failed to demonstrate such reconstitution acted as controls. These patients (mean age 27 years) had chronic HIV infection with low CD4 T-cell counts (mean 82, range 9-160 cells/microL blood). Increased function of the thymus visualized by PET was correlated with phenotypic changes in CD4 and CD8 T cells in the periphery measured by flow cytometry, and with numbers of recent thymic emigrants measured by quantification of the numbers of T-cell receptor excision circles (TRECs) in peripheral cells. RESULTS: In one patient, clear correlations could be drawn between visible activity within the thymus, as measured by increased [F18]fluorodeoxyglucose (FDG) uptake, and regeneration of naïve CD4 (CD45RA/CD62L) T cells, increased numbers of CD4 T cells, controlled viraemia and increased numbers of recent thymic emigrants. A second patient displayed no increase in peripheral CD4 count and no increase in thymic activity. The third patient elected to stop therapy following the 2-month time point. CONCLUSIONS: The use of PET suggests that thymic activity may increase after HAART, indicating that the thymus has the potential to be functional even in HIV-1 infected persons with low CD4 T-cell counts.

Thymic activity in late-stage HIV-1 infected individuals receiving highly active antiretroviral therapy: potential effect of steroid therapy.

OBJECTIVE: Our objective was to monitor the effect of steroid therapy on the thymic output and function of late-stage HIV-1-infected patients undergoing highly active antiretroviral therapy (HAART). DESIGN: The indirect measurement of T cells that have recently emigrated from the thymus as a means of quantifying thymic output, and therefore thymic function, was achieved through use of the polymerase chain reaction-based signal joint T cell receptor rearrangement excision circles (sjTREC) assay. Proliferative capacity and interleukin (IL)-2 and IL-4 production by T cells after antigenic, mitogenic and IL-2 stimulation were also analysed. METHOD: Measurements were made of sjTREC levels in peripheral blood mononuclear cell DNA samples from five HIV-1 infected patients (one on steroid therapy prior to and at the time of sample extraction) receiving HAART. IL-2 and IL-4 production and proliferative capacity were also measured in three patients, including the patient receiving steroids. RESULT: The sjTREC assay gave an extremely weak result for the patient on steroids but, under the same assay conditions, provided clear, positive readings for the four patients not on steroids. Comparison of the patients' cytokine profiles revealed that IL-2 production was generally low or absent in all three patients tested but that IL-4 production was significantly higher in the patient given steroids. Functional potential as revealed by proliferation assays showed very low or absent cellular proliferation. CONCLUSION: The thymic contribution to the restoration of T lymphocyte numbers, particularly during the treatment of HIV-1 infection, may become compromised if thymic inhibitory factors such as steroids are used. Furthermore, the use of steroids may also favour the development of a T helper 2 response, which could prove particularly undesirable during HIV-1 infection.

Use of a high-fidelity simulator to develop testing of the technical performance of novice anaesthetists.

BACKGROUND: We used the Delphi technique to gain a consensus from 26 consultant anaesthetists about technical tasks during general anaesthesia. We then developed a technical scoring system to assess anaesthetists undertaking general anaesthesia with rapid sequence induction. METHODS: We then followed the performance of six novice anaesthetists on five occasions during their first 3 months of training. At each, visit each novice 'anaesthetized' the Human Patient Simulator at Bristol Medical Simulator Centre. For comparison seven post-fellowship anaesthetists were scored on one occasion. RESULTS: Novice scores improved significantly over the 12-week period (P<0.01). A significant difference was also found between the final novice scores and the post-fellowship subjects (P<0.05). CONCLUSIONS: These findings suggest that simulation can be used to observe and quantify technical performance.

Age-associated thymic atrophy is not associated with a deficiency in the CD44(+)CD25(-)CD3(-)CD4(-)CD8(-) thymocyte population.

Age-associated thymic atrophy has been proposed to be due to changes in both the thymic microenvironment and in the intrinsic properties of the early T cell progenitors, the CD44(+)CD25(-)CD3(-)CD4(-)CD8(-) cells. We have purified these cells from the thymus of both old and young mice and demonstrate no age-associated defect in their ability to differentiate into their progeny in vitro when used to reconstitute fetal thymic organ cultures. We also demonstrate that in the presence of anti-IL-7, CD44(+)CD25(-)CD3(-)CD4(-)CD8(-) cells from young mice show reduced thymocyte development in fetal thymic organ cultures compared with controls. Finally we have shown that old mice treated with IL-7 show improved thymopoiesis compared with control groups. The increased thymopoiesis seen in the old animals occurs in the sequential manner which would be anticipated for an agent working directly on the early stages, including the CD44(+)CD25(-)CD3(-)CD4(-)CD8(-) cells.

Gender-related differences in the rates of age associated thymic atrophy.

Age associated thymic atrophy has been shown to be linked to problems with rearrangement of the beta chain of the T cell receptor (TCR) in male mice during the early phases of the intrathymic T cell developmental pathway. In this study, thymic atrophy in female mice was found to occur at a different rate than in male mice. At 9 months of age there was a significantly greater number of cells in the thymus of female mice compared with male mice, with the major difference found in the CD4+CD8+ populations. The thymii of female mice at 9 months of age contained double the number of these cells compared with male mice. Analysis of the CD4+CD8+ cells at 9 months of age demonstrated increased numbers of cells expressing higher levels of CD3 in females compared with males indicating that in females more of these cells were producing successful alphabetaTCR pairings. In F5 transgenic mice comparison of the CD4+CD8+ population revealed no significant difference in their absolute numbers at 9 months of age. These results indicate that the gender differences at this time point were due to fewer permitted divisions prior to the expression of a selectable TCR alpha chain within the CD4+CD8+ populations in male compared with female mice. This gender difference was not due to the action of testosterone and unlikely to be due to differences in the level of oestrogen. The potential mechanisms of this difference may be related to a regulatory feedback of peripheral T cells on the developing thymocyte populations. Such age related changes in the numbers of cells within distinct thymic subpopulations leads to the possibility that the potential repertoire in females is greater than in males later in life.

Both age and gender affect thymic output: more recent thymic migrants in females than males as they age.

The thymus undergoes age-associated involution, with studies showing thymic size decreasing from birth at a rate of approximately 3% per year until middle age, and at a rate of 1% per year thereafter. The aim of this study was to determine the effect of thymic atrophy on T-lymphocyte production by the thymus, and to clarify the ongoing uncertainty regarding gender differences in thymic function. We quantified recent thymic emigrants (RTEs) in blood through the measurement of signal joint T-cell receptor rearrangement excision circles (sjTRECs), and showed that the decline in the number of RTEs in the blood with increasing age is gender-linked. Peripheral blood from females contained significantly higher levels of sjTRECs per CD3+ T cell than blood from males (P = 0.002), despite there being no significant gender difference in the absolute number of CD3+ T cells in the populations analysed (P > 0.10). Our findings suggest better thymic function in females compared with males, providing females with a higher number of recent thymic emigrants for longer periods of life. Such a finding provides a plausible explanation for the immunological gender differences observed in previous studies and possibly, for the general longer life expectancy in females compared with males.

A prospective randomized controlled study of the efficacy of ketamine for postoperative pain relief in children after adenotonsillectomy.

BACKGROUND: Adenotonsillectomy is commonly needed by children with obstructive sleep apnoea syndrome. This population is at high risk of life threatening airway obstruction in the postoperative period. METHODS: Fifty children were studied to test the efficacy of an alternative analgesic to the use of opioids in providing analgesia in the immediate postoperative period. Patients were randomized to receive either 0.1 mg.kg-1 morphine or 0.5 mg.kg-1 ketamine at induction. RESULTS: Ketamine was as effective as morphine with no additional side-effects. CONCLUSIONS: Ketamine is a safe and effective alternative to morphine to provide analgesia in the immediate postoperative period after tonsillectomy.

Effect of CD4(+) and CD8(+) cell depletion on wound healing.

BACKGROUND: Depression of the immune system can result in poor or delayed wound healing. METHODS: Thymectomized rats were depleted of CD4(+) and CD8(+) lymphocytes by intraperitoneal injection of Medical Research Council Oxford (MRC OX)38 antibodies and MRC OX8. Significant depletion was demonstrated throughout the wound healing process by immunofluorescence studies of peripheral blood. Following depletion the rats underwent laparotomy incisions which were allowed to heal for 10 weeks. Differences in healing were demonstrated by analysing the wounds biomechanically by tensiometry to obtain values of ultimate strength, resilience, toughness, maximum extension and elastic constant. RESULTS: Wounds of animals depleted of CD4+ lymphocytes showed a significant decrease in ultimate strength, resilience and toughness. Wounds of animals depleted of CD8(+) lymphocytes showed a significant increase in ultimate strength, resilience and toughness. CONCLUSION: Wounds healed in the absence of T lymphocytes. However, the subsets have an opposing regulatory role, with CD4(+) lymphocytes upregulating and CD8(+) lymphocytes downregulating wound healing. Presented to the Surgical Research Society in Nottingham, UK, 11 July 1997 and published in abstract form as Br J Surg 1997; 84: 1618

Il-7 and not stem cell factor reverses both the increase in apoptosis and the decline in thymopoiesis seen in aged mice.

Thymic atrophy is an age-associated decline in commitment to the T cell lineage considered to be associated with defective TCR beta-chain rearrangement. Both IL-7 and stem cell factor (SCF) have dominant roles at this stage of triple negative (TN) thymocyte development. Because there is no age-associated decrease in the number of CD44(+)CD25(-)CD3(-)CD4(-)CD8(-) cells, this study investigated whether alterations in apoptosis within the TN pathway accounted for diminishing thymocyte numbers with age. Here we show significant age-associated increases in apoptotic TN thymocytes, specifically within CD44(+)CD25(+) and CD44(-)CD25(+) subpopulations, known to be the location of TCR beta-chain rearrangement. IL-7 added to TN cultures established from old mice significantly both reduces apoptosis and increases the percentage of live cells within CD44(+)CD25(+) and CD44(-)CD25(+) subpopulations after 24 h, with prosurvival effects remaining after 5 days. SCF failed to demonstrate prosurvival effects in old or young cultures, and IL-7 and SCF together did not improve upon IL-7 alone. IL-7R expression did not decline with age, ruling out the possibility that the age-associated increase in apoptosis was attributed to reduced IL-7R expression. Compared with PBS, treatment of old mice with IL-7 produced significant increases in live TN cells. By comparison, treatment with SCF failed to increase live TN numbers, and IL-7 and SCF together failed to significantly improve thymopoiesis above that shown by IL-7 alone. Thus, treatment with IL-7 alone can reverse the age-associated defect in TN thymocyte development revealed by in vitro studies to be located at the stages of TCR beta-chain rearrangement.

Erythrocyte transfusion causes immunosuppression after total hip replacement.

A prospective study of 34 patients undergoing total hip replacement was done to determine whether homologous erythrocyte transfusion causes postoperative immunosuppression. In the transfused patient group (14 patients), there was a reduction in CD3+ and CD3+4+ cell numbers at Day 2, returning to preoperative levels by Day 7. In contrast, in the untransfused patient group (20 patients), there was no significant depression in these lymphocyte subgroups at Day 2 and an increase in total lymphocyte, CD3+, CD3+4+, and CD3+4-8- cell numbers at Day 7. In both patient groups there was postoperative leukocytosis, granulocytosis, and monocytosis at Days 2 and 7, with no significant change in postoperative B (CD19+) cell numbers, natural killer cells, or the minor T cell populations of CD3+4+8+ and CD3+25+. The lymphocyte functional test of Candida recall was significantly impaired at Day 7 in the transfused patient group, where Candida recall is a memory T cell response to an antigen extracted from the yeast Candida Albicans. These findings suggest that homologous erythrocyte transfusion after hip replacement surgery causes cell mediated immune suppression. The main clinical implication of the current study is that perioperative homologous erythrocyte transfusion may place patients at greater risk of infectious complications, including infection of the prosthesis.

Thymic involution in aging.

The size of the naive T-cell pool is governed by output from the thymus and not by replication. This pool contributes cells to the activated/memory T-cell pool whose size can be increased through cell multiplication; both pools together constitute the peripheral T-cell pool. Aging is associated with involution of the thymus leading to a reduction in its contribution to the naive T-cell pool; however, despite this diminished thymic output, there is no significant decline in the total number of T cells in the peripheral T-cell pool. There are, however, considerable shifts in the ratios of both pools of cells, with an increase in the number of activated/memory T cells and the accumulation in older individuals of cells that fail to respond to stimuli as efficiently as T cells from younger individuals. Aging is also associated with a greater susceptibility to some infections and some cancers. An understanding of the causal mechanism of thymic involution could lead to the design of a rational therapy to reverse the loss of thymic tissue, renew thymic function, increase thymic output, and potentially improve immune function in aged individuals.