Extracorporeal membrane oxygenation during pregnancy and peripartal. An international retrospective multicenter study.

INTRODUCTION: Extracorporeal Membrane Oxygenation (ECMO) may be used in the setting of pregnancy or the peripartal period, however its utility has not been well-characterized. This study aims to give an overview on the prevalence of peripartel ECMO cases and further assess the indications and outcomes of ECMO in this setting across multiple centers and countries. METHODS: A retrospective, multicenter, international cohort study of pregnant and peripartum ECMO cases was performed. Data were collected from six ECMO centers across three continents over a 10-year period. RESULTS: A total of 60 pregnany/peripartal ECMO cases have been identified. Most frequent indications are acute respiratory distress syndrome (n = 30) and pulmonary embolism (n = 5). Veno-venous ECMO mode was applied more often (77%). ECMO treatment during pregnancy was performed in 17 cases. Maternal and fetal survival was high with 87% (n = 52), respectively 73% (n = 44). CONCLUSIONS: Various emergency scenarios during pregnancy and at time of delivery may require ECMO treatment. Peripartal mortality in a well-resourced setting is rare, however emergencies in the labor room occur and knowledge of available rescue therapy is essential to improve outcome. Obstetricians and obstetric anesthesiologists should be aware of the availability of ECMO resource at their hospital or region to ensure immediate contact when needed.

Targeted Temperature Management After In-Hospital Cardiac Arrest: An Ancillary Analysis of Targeted Temperature Management for Cardiac Arrest With Nonshockable Rhythm Trial Data.

BACKGROUND: Targeted temperature management (TTM) currently is the only treatment with demonstrated efficacy in attenuating the harmful effects on the brain of ischemia-reperfusion injury after cardiac arrest. However, whether TTM is beneficial in the subset of patients with in-hospital cardiac arrest (IHCA) remains unclear. RESEARCH QUESTION: Is TTM at 33 °C associated with better neurological outcomes after IHCA in a nonshockable rhythm compared with targeted normothermia (TN; 37 °C)? STUDY DESIGN AND METHODS: We performed a post hoc analysis of data from the published Targeted Temperature Management for Cardiac Arrest with Nonshockable Rhythm randomized controlled trial in 584 patients. We included the 159 patients with IHCA; 73 were randomized to 33 °C treatment and 86 were randomized to 37 °C treatment. The primary outcome was survival with a good neurologic outcome (cerebral performance category [CPC] score of 1 or 2) on day 90. Mixed multivariate adjusted logistic regression analysis was performed to determine whether survival with CPC score of 1 or 2 on day 90 was associated with type of temperature management after adjustment on baseline characteristics not balanced by randomization. RESULTS: Compared with TN for 48 h, hypothermia at 33 °C for 24 h was associated with a higher percentage of patients who were alive with good neurologic outcomes on day 90 (16.4% vs 5.8%; P = .03). Day 90 mortality was not significantly different between the two groups (68.5% vs 76.7%; P = .24). By mixed multivariate analysis adjusted by Cardiac Arrest Hospital Prognosis score and circulatory shock status, hypothermia was associated significantly with good day 90 neurologic outcomes (OR, 2.40 [95% CI, 1.17-13.03]; P = .03). INTERPRETATION: Hypothermia at 33 °C was associated with better day 90 neurologic outcomes after IHCA in a nonshockable rhythm compared with TN. However, the limited sample size resulted in wide CIs. Further studies of patients after cardiac arrest resulting from any cause, including IHCA, are needed.

Cerebrovascular Autoregulation Monitoring in the Management of Adult Severe Traumatic Brain Injury: A Delphi Consensus of Clinicians.

BACKGROUND: Several methods have been proposed to measure cerebrovascular autoregulation (CA) in traumatic brain injury (TBI), but the lack of a gold standard and the absence of prospective clinical data on risks, impact on care and outcomes of implementation of CA-guided management lead to uncertainty. AIM: To formulate statements using a Delphi consensus approach employing a group of expert clinicians, that reflect current knowledge of CA, aspects that can be implemented in TBI management and CA research priorities. METHODS: A group of 25 international academic experts with clinical expertise in the management of adult severe TBI patients participated in this consensus process. Seventy-seven statements and multiple-choice questions were submitted to the group in two online surveys, followed by a face-to-face meeting and a third online survey. Participants received feedback on average scores and the rationale for resubmission or rephrasing of statements. Consensus on a statement was defined as agreement of more than 75% of participants. RESULTS: Consensus amongst participants was achieved on the importance of CA status in adult severe TBI pathophysiology, the dynamic non-binary nature of CA impairment, its association with outcome and the inadvisability of employing universal and absolute cerebral perfusion pressure targets. Consensus could not be reached on the accuracy, reliability and validation of any current CA assessment method. There was also no consensus on how to implement CA information in clinical management protocols, reflecting insufficient clinical evidence. CONCLUSION: The Delphi process resulted in 25 consensus statements addressing the pathophysiology of impaired CA, and its impact on cerebral perfusion pressure targets and outcome. A research agenda was proposed emphasizing the need for better validated CA assessment methods as well as the focused investigation of the application of CA-guided management in clinical care using prospective safety, feasibility and efficacy studies.

Correction to: Serum sodium and intracranial pressure changes after desmopressin therapy in severe traumatic brain injury patients: a multi-centre cohort study.

Following publication of the original article [1], we were notified that the collaborators' names part of the "The TBI Collaborative" group has not been indexed in Pubmed. Below the collaborators names full list.

Serum sodium and intracranial pressure changes after desmopressin therapy in severe traumatic brain injury patients: a multi-centre cohort study.

BACKGROUND: In traumatic brain injury (TBI) patients desmopressin administration may induce rapid decreases in serum sodium and increase intracranial pressure (ICP). AIM: In an international multi-centre study, we aimed to report changes in serum sodium and ICP after desmopressin administration in TBI patients. METHODS: We obtained data from 14 neurotrauma ICUs in Europe, Australia and UK for severe TBI patients (GCS ≤ 8) requiring ICP monitoring. We identified patients who received any desmopressin and recorded daily dose, 6-hourly serum sodium, and 6-hourly ICP. RESULTS: We studied 262 severe TBI patients. Of these, 39 patients (14.9%) received desmopressin. Median length of treatment with desmopressin was 1 [1-3] day and daily intravenous dose varied between centres from 0.125 to 10 mcg. The median hourly rate of decrease in serum sodium was low (- 0.1 [- 0.2 to 0.0] mmol/L/h) with a median period of decrease of 36 h. The proportion of 6-h periods in which the rate of natremia correction exceeded 0.5 mmol/L/h or 1 mmol/L/h was low, at 8% and 3%, respectively, and ICPs remained stable. After adjusting for IMPACT score and injury severity score, desmopressin administration was independently associated with increased 60-day mortality [HR of 1.83 (1.05-3.24) (p = 0.03)]. CONCLUSIONS: In severe TBI, desmopressin administration, potentially representing instances of diabetes insipidus is common and is independently associated with increased mortality. Desmopressin doses vary markedly among ICUs; however, the associated decrease in natremia rarely exceeds recommended rates and median ICP values remain unchanged. These findings support the notion that desmopressin therapy is safe.

Elderly versus nonelderly patients with invasive aspergillosis in the ICU: a comparison and risk factor analysis for mortality from the AspICU cohort.

Data regarding the epidemiology and diagnosis of invasive aspergillosis in the critically ill population are limited, with data regarding elderly patients (≥75 years old) even scarcer. We aimed to further compare the epidemiology, characteristics and outcome of elderly versus nonelderly critically ill patients with invasive aspergillosis (IA) Prospective, international, multicenter observational study (AspICU) including adult intensive care unit (ICU) patients, with a culture and/or direct examination and/or histopathological sample positive for Aspergillus spp. at any site. We compared clinical characteristics and outcome of IA in ICU patients using two different diagnostic algorithms. Elderly and nonelderly ICU patients with IA differed in a number of characteristics, including comorbidities, clinical features of the disease, mycology testing, and radiological findings. No difference regarding mortality was found. According to the clinical algorithm, elderly patients were more likely to be diagnosed with putative IA. Elderly patients had less diagnostic radiological findings and when these findings were present they were detected late in the disease course. The comparison between elderly survivors and nonsurvivors demonstrated differences in clinical characteristics of the disease, affected sites and supportive therapy needed. All patients who were diagnosed with proven IA died. Increased vigilance combined with active search for mycological laboratory evidence and radiological confirmation are necessary for the timely diagnosis of IA in the elderly patient subset. Although elderly state per se is not a particular risk factor for mortality, a high SOFA score and the decision not to administer antifungal therapy may have an impact on survival of elderly patients.

Management strategies in venovenous extracorporeal membrane oxygenation: a retrospective comparison from five European centres.

OBJECTIVE: To investigate contemporary clinical practice in the management of venovenous (VV) extracorporeal membrane oxygenation (ECMO) in critically ill patients with bacterial pneumonia. METHODS: In this multicentre retrospective study, 48 patients with severe respiratory failure due to bacterial pneumonia receiving VV ECMO therapy in five experienced European ECMO centres were included. Ventilator and ECMO settings were analysed. RESULTS: Ventilator settings showed great variability between participating centres, particularly relating to positive end-expiratory pressure, peak inspiratory pressure and driving pressure. Different strategies in cannulation, ECMO setting and weaning procedures were also observed. CONCLUSION: There is great diversity in management modalities for ventilator and ECMO settings for patients with bacterial pneumonia. Our study emphasises the lack of clinical consensus in VV ECMO management.

Drivers and impact of antifungal therapy in critically ill patients with Aspergillus-positive respiratory tract cultures.

Invasive pulmonary aspergillosis (IPA) is an increasingly recognised problem in critically ill patients. Little is known about how intensivists react to an Aspergillus-positive respiratory sample or the efficacy of antifungal therapy (AFT). This study aimed to identify drivers of AFT prescription and diagnostic workup in patients with Aspergillus isolation in respiratory specimens as well as the impact of AFT in these patients. ICU patients with an Aspergillus-positive respiratory sample from the database of a previous observational, multicentre study were analysed. Cases were classified as proven/putative IPA or Aspergillus colonisation. Demographic, microbiological, diagnostic and therapeutic data were collected. Outcome was recorded 12 weeks after Aspergillus isolation. Patients with putative/proven IPA were more likely to receive AFT than colonised patients (78.7% vs. 25.5%; P <0.001). Patients with host factors for invasive fungal disease were more likely to receive AFT (72.5% vs. 37.4%) as were those with multiorgan failure (SOFA score >7) (68.4% vs. 36.9%) (both P <0.001). Once adjusted for disease severity, initiation of AFT did not alter the odds of survival (HR = 1.40, 95% CI 0.89-2.21). Likewise, treatment within 48 h following diagnosis did not change the clinical outcome (75.7% vs. 61.4%; P = 0.63). Treatment decisions appear to be based on diagnostic criteria and underlying disease severity at the time of Aspergillus isolation. IPA in this population has a dire prognosis and AFT is not associated with reduced mortality. This may be explained by delayed diagnosis and an often inevitable death due to advanced multiorgan failure.

Early-goal directed therapy for septic shock: is it the end?

Three randomized clinical trials have recently provided data on the lack of effectiveness of "early-goal directed therapy" (EGDT) (i.e. optimization of tissue oxygenation in the first 6 hours since sepsis diagnosis using different therapeutic interventions based on the assessment of the central venous oxygen saturation to titrate such interventions) in the initial management of patients with septic shock. In a first trial including 31 US hospitals (the ProCESS study, N.=1341), three different therapeutic strategies (EGDT vs. protocol-based therapy vs. usual care) were compared and no difference in the primary endpoint (60-day mortality) was found (EGDT 21%, protocol-based therapy 18% and usual care 19%). No significant difference in death by 90 days or in other secondary outcomes, including serious adverse events, was found, as well. A second trial (ARISE, N.=1600), mostly conducted in Australia and New Zealand, randomized patients to EGDT or usual care. Ninety-day mortality was similar between groups (19% vs. 19%, respectively; P=0.90) and no other differences in secondary endpoints were recorded between the two groups. A third study (ProMISe, N.=1260) included patients in 56 hospitals across England, randomly assigned to EGDT or usual care. By 90 days, mortality was similar between groups (29% vs. 29%, respectively; P=0.90). Moreover, EGDT significantly increased costs and was associated with a longer hospital length of stay. We discussed some issues related to the differences between these studies and the pivotal paper from Rivers et al. and how EGDT should be still considered in the treatment of sepsis.

How to monitor the brain in septic patients?

Brain injury is frequently observed after sepsis and may be primarily related to the direct effects of the septic insult on the brain (e.g., brain edema, ischemia, seizures) or to secondary/indirect injuries (e.g., hypotension, hypoxemia, hypocapnia, hyperglycemia). Management of brain injury in septic patients is first focused to exclude structural intracranial complications (e.g., ischemic/hemorrhagic stroke) and possible confounders (e.g., electrolyte alterations or metabolic disorders, such as dysglycemia). Sepsis-associated brain dysfunction is frequently a heterogeneous syndrome. Despite increasing understanding of main pathophysiologic determinants, therapy is essentially limited to protect the brain against further cerebral damage, by way of "simple" therapeutic manipulations of cerebral perfusion and oxygenation and by avoiding over-sedation. Non-invasive monitoring of cerebral perfusion and oxygenation with transcranial Doppler (TCD) and near-infrared spectroscopy (NIRS) is feasible in septic patients. Electroencephalography (EEG) allows detection of sepsis-related seizures and holds promise also as sedation monitoring. Brain CT-scan detects intra-cerebral structural lesions, while magnetic resonance imaging (MRI) provides important insights into primary mechanisms of sepsis-related direct brain injury, (e.g., cytotoxic vs. vasogenic edema) and the development of posterior reversible encephalopathy. Together with EEG and evoked potentials (EP), MRI is also important for coma prognostication. Emerging clinical evidence suggests monitoring of the brain in septic patients can be implemented in the ICU. The objective of this review was to summarize recent clinical data about the role of brain monitoring - including TCD, NIRS, EEG, EP, CT, and MRI - in patients with sepsis and to illustrate its potential utility for the diagnosis, management and prognostication.

Brain injury after cardiac surgery.

In patients undergoing cardiac surgery, postoperative brain injury significantly contributes to increase morbidity and mortality and has negative consequences on quality of life and costs. Moreover, over the past years, compelling medical and technological improvements have allowed an even older patients' population, with several comorbidities, to be treated with cardiac surgery; however, the risk of brain injury after such interventions is also increased in these patients. With the aim of improving post-operative neurological outcome, a variety of neuromonitoring methods and devices have been introduced in clinical practice. These techniques allow the assessment of a number of parameters, such as cerebral blood flow, brain embolic events, cerebral cortical activity, depth of anesthesia and brain oxygenation. Some of them have been used to optimize the hemodynamic management of such patients and to select specific therapeutic interventions. Also, various pharmacological and non-pharmacological approaches have been proposed to minimize the incidence of brain injury in this setting. In this review we describe the risk factors and mechanisms of cerebral injury after cardiac surgery and focus on monitoring techniques and clinical strategies that could help clinicians to minimize the incidence of brain injury.

Elevated ß-lactam concentrations associated with neurological deterioration in ICU septic patients.

BACKGROUND: Although ß-lactams are considered to have a safe therapeutic profile, neurotoxicity has been reported. The aim of this study was to assess the association between ß-lactam concentrations and neurological alterations in septic ICU patients. METHODS: Retrospective study on all ICU patients who were treated with meropenem (MEM), piperacillin-tazobactam (TZP) or ceftazidime/cefepime (CEF) and in whom at least one ß-lactam trough concentration (C min) was determined. Drug levels were measured using high-performance liquid chromatography; C min was normalized to the clinical breakpoint of Pseudomonas aeruginosa (as determined by EUCAST) for each drug (C min/MIC). Changes in neurological status were evaluated using changes in the neurological sequential organ failure assessment score (ΔnSOFA) using the formula: ΔnSOFA = nSOFA(day of TDM) - nSOFA(ICU admission). Worsening neurological status (NWS) was defined as a ΔnSOFA ≥ 1 for an nSOFA on admission of 0-2. RESULTS: We collected 262 C min in 199 patients (130 MEM, 85 TZP, 47 CEF). Median APACHE II score and GCS on admission were 17 and 15, respectively. Overall ICU mortality was 27 %. There were no differences in the occurrence of NWS between antibiotics (39% for MEM, 32% for TZP and 35% for CEF). The occurrence of NWS increased with increasing C min/MIC ranges (P = 0.008); this correlation was found for TZP (P = 0.05) and MEM (P = 0.01), but not for CEF. C min/MIC was an independent predictive factor for NWS (OR 1.12 [1.04-1.20]). CONCLUSION: We found a correlation between high ß-lactam trough concentrations and increased occurrence of neurological deterioration in septic ICU patients. Although our data cannot determine causality, monitoring of ß-lactam levels should be considered when deterioration of neurological status occurs during critical illness.

Colloids for fluid resuscitation: what is their role in patients with shock?

BACKGROUND: Fluid therapy is widely used in critically ill patients to restore effective intravascular volume and improve organ perfusion. Recent studies have questioned the administration of colloid-based solutions, especially if containing hydroxyethyl starch (HES), in different ICU populations; however, there is still uncertainty on the use of colloids as initial fluid therapy for early resuscitation. METHODS: The aim of this study was to investigate the effect of two different resuscitation fluid strategies on the mortality of patients with shock. In a multicentric (57 ICUs), controlled, open-label trial (from February 2003 to August 2012), the authors randomized patients with signs of acute hypovolemia, defined by the combination of hypotension, evidence of low filling pressures or cardiac index and at least two signs of tissue hypoperfusion (such as altered consciousness, mottled skin, oliguria, lactate levels > 2 mmol/L), to received either a colloid- or crystalloid-based therapy. Both cohorts received maintenance fluids consisting in isotonic crystalloids and albumin in case of severe hypoalbuminemia (<2 g/dL). Exclusion criteria included previous fluid therapy, pregnancy, brain death, extended burns, chronic hemodialysis or liver disease, known coagulopathy, acute anaphylaxis, dehydration and hypotension due to sedative drugs. RESULTS: Among the 6498 eligible patients, 2857 were eventually randomized in one of the two groups. The 28-day mortality was 25.4% in the colloid and 27.0% in the crystalloid group (P=0.26). In the sub-group analysis, similar mortality rates were reported for shock due to hypovolemia, sepsis or trauma. Also, the use of continuous renal replacement therapy was similar between groups (11.0% vs. 12.5%, P=0.19). There were more days alive without mechanical ventilation or vasopressors during the first 7 and 28 days and a lower 90-day mortality in the colloid group. CONCLUSION: These data suggest that mortality was not increased and probably decreased with the use of colloids in different forms of shock requiring early fluid resuscitation. We discussed herein some methodological issues that may explain the discrepancies of this trial with the other studies developed in the same field.

Glucocorticoid administration in sepsis and septic shock: time for a paradigm change?

The use of corticosteroids in patients with septic shock remains controversial. Questions remain regarding the more appropriate dose, the optimal timing to initiate therapy, the selection of patients who will benefit most from the treatment and the exact mechanisms involved in their effectiveness. Recent studies have highlighted that, in critically ill patients, corticosteroid metabolism was reduced and associated with high circulating cortisol levels. Hence the required doses of hydrocortisone may be lower than the currently recommended doses in septic shock (i.e. 200 mg/day). However, altered expression and/or function of corticosteroid receptors may still suggest that higher hydrocortisone doses are necessary to overcome this so-called "steroid-resistance". In this article, we summarized these recent concepts and discussed how they could influence the administration of corticosteroids in such patients.

Broad-spectrum ß-lactams in obese non-critically ill patients.

OBJECTIVES: Obesity may alter the pharmacokinetics of ß-lactams. The goal of this study was to evaluate if and why serum concentrations are inadequate when standard ß-lactam regimens are administered to obese, non-critically ill patients. SUBJECTS AND METHODS: During first year, we consecutively included infected, obese patients (body mass index (BMI) ⩾30 kg m(-2)) who received meropenem (MEM), piperacillin-tazobactam (TZP) or cefepime/ceftazidime (CEF). Patients with severe sepsis or septic shock, or those hospitalized in the intensive care unit were excluded. Serum drug concentrations were measured twice during the elimination phase by high-performance liquid chromatography. We evaluated whether free or total drug concentrations were >1 time (fT>minimal inhibition concentration (MIC)) or >4 times (T>4MIC) the clinical breakpoints for Pseudomonas aeruginosa during optimal periods of time: ⩾40% for MEM, ⩾50% for TZP and ⩾70% for CEF. RESULTS: We included 56 patients (median BMI: 36 kg m(-2)): 14 received MEM, 31 TZP and 11 CEF. The percentage of patients who attained target fT>MIC and T>4MIC were 93% and 21% for MEM, 68% and 19% for TZP, and 73% and 18% for CEF, respectively. High creatinine clearance (107 (range: 6-398) ml min(-1)) was the only risk factor in univariate and multivariate analyses to predict insufficient serum concentrations. CONCLUSIONS: In obese, non-critically ill patients, standard drug regimens of TZP and CEF resulted in insufficient drug concentrations to treat infections due to less susceptible bacteria. Augmented renal clearance was responsible for these low serum concentrations. New dosage regimens need to be explored in this patient population (EUDRA-CT: 2011-004239-29).

How to target temperature after cardiac arrest: insights from a randomized clinical trial.

Implementation of treatments able to improve survival and neurological recovery of cardiac arrest (CA) survivors is a major clinical challenge. More than ten years ago, two pivotal trials showed that application of therapeutic hypothermia (TH, 32-34 °C) to patients resuscitated from an out-of-hospital CA (OHCA) with an initial shockable rhythm significantly ameliorated their outcome. Since then, TH has been used also for non-shockable rhythms and for in-hospital CA to some extent, even if the quality of evidence supporting TH in such situations remained very low. The objective of this randomized, controlled, multicenter study (named "Targeted Temperature Management" TTM study) was to compare two different strategies of temperature control after CA; patients were randomized to be treated either at 33 °C or at 36 °C for 24 hours, while fever was accurately avoided for the first 3 days since randomization. Inclusion criteria were: Glasgow Coma Score <8, presumed cardiac origin of arrest, randomization occurring within the first 4 hours from the return of spontaneous circulation. Patients were excluded if they had an unwitnessed arrest with asystole as the initial rhythm, suspected or known acute intracranial hemorrhage or stroke, and a body temperature of less than 30 °C. A specific algorithm was used to decide for withdrawal of care in patients remaining comatose after 72 hours since normothermia was achieved. The primary outcome was 6-month mortality. After the enrollment of 939 patients, the authors did not find any significant difference between groups in primary outcome (235/473 [50%] and 225/466 [48%] of patients died in 33 °C and 36 °C group, respectively; HR for death if in the 33 °C group, 1.06 [95% CI 0.89 to 1.28; P=0.51]). Similarly, the analysis of the composite outcome of death or poor neurologic function yielded similar results between the two groups. This is the largest study evaluating the effects of two different strategies of temperature management after CA. Some important concerns have been raised on the real benefit of keeping CA patients at 33 °C and major changes in clinical practice are expected. We discussed herein the main differences with previous randomized trials and tried to identify possible explanations for these findings.

Esmolol for septic shock: more than just heart rate control?

Excessive adrenergic stimulation may be associated with several adverse events and contribute to increase mortality in critically ill septic patients. Few clinical data exist on the effects of adrenergic blockade in this setting. The objective of this study was to investigate the effect of a short acting b-blocker (esmolol) in septic shock patients. In a single-center, controlled, open-label, phase 2 trial (from November 2010 to July 2012), Morelli et al. randomized patients with a need of norepinephrine to maintain a mean arterial pressure above 65 mmHg to receive either esmolol or standard of care. Patients were included if, after 24 hours of initial resuscitation, hypovolemia was excluded (wedge pressure ≥12 mmHg or central venous pressure ≥8 mmHg) and heart rate was above 95 bpm. Patients were excluded if they were younger than 18 years, had previous b-blockers therapy, cardiac index was ≤2.2 L/min/m² with wedge pressure >18 mmHg, were diagnosed with significant cardiac valvular diseases or were pregnant. The primary outcome was the reduction in heart rate between 80 and 94 bpm over a 96-hr period. Secondary outcomes included norepinephrine requirement, hemodynamic changes, organ function, adverse events and 28-day mortality. A total of 154 patients, 77 for each group, were enrolled. Esmolol was more effective than standard treatment to reduce heart rate within target limits; also, b-blocker therapy was associated with an increased stroke volume and left ventricular work index when compared to the control group. These favorable hemodynamic effects were associated with a better control of lactate levels, a higher reduction in norepinephrine and fluids requirement. Mortality was 49.4% in the esmolol group and 80.5% in the control group (P<0.01). This is the first study showing an improvement in cardiac function and 28-day mortality in septic patients adding b-blockers to standard therapy. We discussed several statistical and methodological limitations that may influence the generability of these results.