Dabigatran: A new oral anticoagulant. Guidelines to follow in oral surgery procedures. A systematic review of the literature.

BACKGROUND: Dabigatran is a newly commercialized drug that is replacing other anticoagulants in the prevention of venous thromboembolism, stroke and systemic arterial valve embolism. It acts directly on thrombin presenting in a dynamic and predictable way, which does not require monitoring these patients. Therefore, we consider the need to assess whether their use increases the risk of bleeding involved before any dental treatment. MATERIAL AND METHODS: We performed a systematic review with a bibliographic search in PubMed/Medline along with the Cochrane Library. We excluded articles dealing with all anticoagulants other than dabigatran, and works about surgical treatments in anatomical locations other than the oral cavity. RESULTS: We included a total of 13 papers of which 1 was a randomized clinical trial, 9 narrative literature reviews, 1 case series, 2 clinical cases and 1 expert opinion. Because we did not obtain any properly designed clinical trials, we were unable to conduct a meta-analysis. CONCLUSIONS: Currently, there is no consensus on the procedure to be followed in patients taking dabigatran. However, all authors agree to treat each case individually in accordance to the risk of embolism, postoperative bleeding and renal function. Also, it is necessary to perform minimally invasive interventions, and take the appropriate local anti-hemolytic measures.

Altered visual sensory fusion in children with reading difficulties.

Reading is a multi-sensory and multi-cognitive task, and its difficulties (e.g., dyslexia) are not a unitary disorder. There are probably a variety of manifestations that relate to the actual site of impairment. A randomized, pre-test/post-test nonequivalent-groups design was conducted over 4 months with three groups aged between 6 and 8 years. One group comprised 76 participants (34 boys, 42 girls) with reading difficulties and altered sensory fusion (RD+ASF), a second group was made up of 123 students (59 boys, 64 girls) with reading difficulties but without altered sensory fusion (RD), and a third group comprised 81 participants (39 boys, 42 girls) who were young readers (RL) without reading delay, paired with the RD group on reading level. The experimental groups received intervention in the skills of control, stimulus recognition, and phonological awareness during a 4-month period. Both pre-test and post-test measures of errors in reading mechanics and reading routes (word and pseudo-word) were obtained. Poorer results in mechanics and reading routes of the RD+ASF group suggest that the effectiveness of the intervention depended on the characteristics of the groups and on the presence of sensory fusion deficits in the RD students.

Development of lymphoproliferative disease after liver transplantation.

INTRODUCTION: Malignancies are a serious long-term complication among liver transplant recipients, with an overall incidence of 4.5%-15%. Posttransplantation lymphoproliferative disease (PTLD) is one of the leading causes of late death. Its development is related to complex interactions between immunosuppressive drugs and environmental agents. The aim of this study was to analyze risk factors for PTLD and survival after orthotopic liver transplantation (OLT) compared with solid tumors. PATIENTS AND METHODS: We undertook a retrospective review of the clinical histories of adult patients who underwent OLT between July 1986 and February 2001, and who had been followed until 2005. This study comprised 528 adult recipients who survived more than 2 months after OLT. We excluded pediatric, partial-organ, and multiorgan recipients. RESULTS: No differences were observed concerning gender, viral etiology of hepatitis, calcineurin inhibitor regimen, or steroid maintenance period. Treated acute rejection episodes accounted for 53.3% of patients who developed PTLD compared with 47.3% in the control group (P = .787). Patients with solid tumors were older at the time of diagnosis than those with PTLD (57.5 +/- 8.13 years vs 48.8 +/- 13.9; P = .002). The overall mortality rate for PTLD was 55.5%, which did not differ significantly from solid tumors. CONCLUSIONS: PTLD develops in younger patients after OLT. Various immunosuppressive regimens do not seem to influence the incidence of PTLD or other solid tumors.

Changes in brain oxidative metabolism induced by water maze training.

Although the hippocampus has been shown to be essential for spatial memory, the contribution of associated brain regions is not well established. Wistar rats were trained to find a hidden escape platform in the water maze during eight days. Following training, the oxidative metabolism in different brain regions was evaluated using cytochrome oxidase histochemistry. Metabolic activations were found in the prelimbic cortex, cornu ammonis (CA) 1 subfield of the dorsal hippocampus and the anterior thalamic nuclei, relative to yoked swim controls and naïve rats. In addition, many cross-correlations in brain metabolism were observed among the latter regions. These results support the implication of a hippocampal-prefrontal-thalamic system to spatial memory in rats.

Virological analysis in the diagnosis of sudden children death: a medico-legal approach.

Infections are considered to be an important cause of unexpected death in children. It has also been assumed that respiratory viruses are involved in the genesis of sudden infant death syndrome (SIDS). The Spanish National Institute of Toxicology and Forensic Sciences act as the forensic reference centre for Spain. We analyse the experience of this centre in the virological study of 64 cases of sudden children death where viral serology, virological cultures, herpesviruses polymerase chain reaction (PCR) and electron microscopy were performed. According to pathological findings, death could only be attributed to an adenovirus infection in one amygdalitis with upper airways stenosis and asphyxia. Human herpes virus 6 (HHV-6) was detected by PCR in one case with pathological findings characteristic of SIDS. Recent infection by respiratory syncytial virus (RSV), Epstein-Barr virus (EBV) and cytomegalovirus (CMV) were also detected. Meanwhile, 85.9% of the cases yielded negative viral results. Twenty-eight infants were finally categorised as SIDS. Pathological findings of infection were detected in 12 patients despite the negativity of viral analyses. Although viral infection is an uncommon cause of sudden children death, a complete microbiological investigation will help to solve the puzzle of SIDS. Definitive guidelines for microbiological analyses need to be updated whilst new pathogens are discovered or new techniques are implemented in order to clarify unsolved cases.

The use of piezoelectric surgery to lateralize the inferior alveolar nerve with simultaneous implant placement and immediate buccal cortical bone repositioning: a prospective clinical study.

A prospective study was conducted to assess a variation of inferior alveolar nerve (IAN) lateralization. This study included 13 patients. An osteotomy was made with a piezoelectric device, and the IAN bundle was moved buccally. Dental implants were then inserted medial to the nerve bundle, and the inner surface of the buccal cortical bone plate was shaped to reduce its thickness. Finally, the bone plate was repositioned to restore the original shape and contour of the mandible. Neurosensory examinations of the lower lip and chin were performed using three tests: light touch, pain, and two-point discrimination. Three months after surgery, the function of the IAN was judged to be completely restored at 11 of the 13 surgical sites. Differences in the tests comparing the operated and non-operated sides were not significant. No implants were lost, and all patients were satisfied with the result. Although IAN lateralization in conjunction with dental implant placement is rarely indicated, the use of a piezoelectric device to perform a buccal osteotomy with final repositioning of the buccal cortical plate over the bony defect contributes to the recovery of the contour and shape of the mandible, without impairment of IAN function.

Mutation rates at Y chromosome specific microsatellites.

A collaborative work was carried out by the Spanish and Portuguese ISFG Working Group (GEP-ISFG) to estimate Y-STR mutation rates. Seventeen Y chromosome STR loci (DYS19, DYS385, DYS389I and II, DYS390, DYS391, DYS392, DYS393, DYS437, DYS438, DYS439, DYS460, DYS461, DYS635 [GATA C4], GATA H4, and GATA A10) were analyzed in a sample of 3,026 father/son pairs. Among 27,029 allele transfers, 54 mutations were observed, with an overall mutation rate across the 17 loci of 1.998 x 10(-3) (95% CI, 1.501 x 10(-3) to 2.606 x 10(-3)). With just one exception, all of the mutations were single-step, and they were observed only once per gametogenesis. Repeat gains were more frequent than losses, longer alleles were found to be more mutable, and the mutation rate seemed to increase with the father's age. Hum Mutat 26(6), 520-528, 2005. (c) 2005 Wiley-Liss, Inc.

Molecular characterisation of Trypanosoma rangeli strains isolated from Rhodnius ecuadoriensis in Peru, R. colombiensis in Colombia and R. pallescens in Panama, supports a co-evolutionary association between parasites and vectors.

We present data on the molecular characterisation of strains of Trypanosoma rangeli isolated from naturally infected Rhodnius ecuadoriensis in Peru, from Rhodnius colombiensis, Rhodnius pallescens and Rhodnius prolixus in Colombia, and from Rhodnius pallescens in Panama. Strain characterisation involved a duplex PCR with S35/S36/KP1L primers. Mini-exon gene analysis was also carried out using TrINT-1/TrINT-2 oligonucleotides. kDNA and mini-exon amplification indicated dimorphism within both DNA sequences: (i) KP1, KP2 and KP3 or (ii) KP2 and KP3 products for kDNA, and 380 bp or 340 bp products for the mini-exon. All T. rangeli strains isolated from R. prolixus presented KP1, KP2 and KP3 products with the 340 bp mini-exon product. By contrast, all T. rangeli strains isolated from R. ecuadoriensis, R. pallescens and R. colombiensis, presented profiles with KP2 and KP3 kDNA products and the 380 bp mini-exon product. Combined with other studies, these results provide evidence of co-evolution of T. rangeli strains associated with different Rhodnius species groups east and west of the Andean mountains.

Latex agglutination for bacterial antigens and meningococcus PCR: two useful tools in legal sudden deaths.

Bacterial infections are considered to be a major cause of sudden deaths. The recognition of infections caused by Neisseria meningitidis is an essential duty of medicolegal offices due to the risk of secondary cases. Since other microorganisms, such as Haemophilus influenzae and Streptococcus pneumoniae, are also involved in infectious sudden deaths, the identification of the pathogen responsible for death is essential in order to establish a positive diagnosis while also preventing secondary meningococcal cases. However, because of the unreliability of culture methods used for autopsy specimens and the fragile nature of the microorganisms, other techniques were used. In this study, the detection of specific antigens of N. meningitidis (serogroups A, B, C, Y and W135), H. influenzae type b, S. pneumoniae and Group B Streptococcus was undertaken in 40 samples from sudden death cases in legal procedures with a latex agglutination test. In addition, a meningococcus polymerase chain reaction (PCR) assay (ctrA, crgA and siaD genes) was also used as a corroboration method for positive N. meningitidis agglutinations. Eleven cases of sudden death were confirmed to be due to meningococcus while one case was confirmed to have been caused by H. influenzae type b fulminant epiglottitis. Rapid laboratory diagnosis of meningococcal infection allowed contacts management and notification to the health authorities. From the point of view of the authors, forensic diagnosis of unascertained deaths should include latex agglutination and meningococcus PCR when a fulminant infection by N. meningitidis or H. influenzae is suspected as well as in deaths where the cause is unclear.

[Comparison between the 1993 and 2002 Guidelines of the Spanish Society of Pulmonology and Thoracic Surgery (SEPAR) for identifying respiratory events in polysomnography tests]. / Comparación entre las normativas de la SEPAR de 1993 y 2002 en la lectura de los eventos respiratorios de las mismas polisomnografías.

OBJECTIVE: To compare the results of applying both the 1993 and 2002 guidelines of the Spanish Society of Pulmonology and Thoracic Surgery (SEPAR) to identify respiratory events during nighttime polysomnography tests. PATIENTS AND METHODS: One hundred twenty consecutive patients with medium to high suspicion of sleep apnea-hypopnea syndrome (SAHS) were included in the study. The 1993 guidelines recommended the use of a thermistor and the evaluation of only apneas and hypopneas. The 2002 guidelines, on the other hand, recommended the use of a thermistor, nasal pressure cannula, and thoracoabdominal bands so that respiratory effort related to arousals could be studied along with apneas and hypopneas. In our study we did not use an esophageal pressure catheter. We calculated the apnea index, hypopnea index, and apnea-hypopnea index (AHI) and determined the number of patients who would be diagnosed with SAHS (AH I > or =10) and the number for whom initiation of continuous positive airway pressure treatment would be recommended (AHI > or =30) according to the 2 sets of guidelines. RESULTS: Polysomnographic tests were valid for 118 of the 120 patients (80% men). The mean (SD) age was 51 (11.6) years and the mean body mass index 31.2 (4.3). Using the 1993 guidelines, the AHI was less than 10 in 25 patients, between 20 and 29 in 38, and 30 or more in 50. In the group overall, mean apnea and hypopnea indices and AHI were all significantly higher with the 2002 guidelines than with the 1993 criteria. With the 1993 criteria, the mean AHI was 33.16 and with 2002 criteria, 45.02 (P<.05). Sixty-four percent of the studies considered normal according to the 1993 SEPAR guidelines were considered apneic according to the 2002 guidelines. Of the patients considered not to need continuous positive airway pressure according to the 1993 SEPAR guidelines, 47.61% did need therapy according to the 2002 guidelines. CONCLUSIONS: There are significant differences in AHI, and in both apnea and hypopnea indices depending on whether the 1993 or the 2002 SEPAR guidelines are applied.

The codon 213 of the 11beta-hydroxysteroid dehydrogenase type 2 gene is a hot spot for mutations in apparent mineralocorticoid excess.

In the kidney, the 11beta-hydroxysteroid dehydrogenase type 2 enzyme (11betaHSD2) inactivates glucocorticoids to their inactive ketoforms and thus prevents endogenous glucocorticoids from occupying the nonselective mineralocorticoid receptor in epithelial tissues. Several mutations have been described in the 11betaHSD2 gene in the congenital syndrome of apparent mineralocorticoid excess. These mutations generate partially or completely inactive 11betaHSD2 enzymes. In the present work, we describe an already known mutation in a new patient affected by apparent mineralocorticoid excess, which results in an arginine-to-cysteine mutation (R213C) in the 11betaHSD2 enzyme. This mutation has been found in two other independent families. In vitro expression studies of this mutant provide evidence that the mutant protein is normally expressed, but its activity is abolished. The CGC-to-TGC (C-toT) transition at codon 213 can be considered a typical CpG-consequence mutation. The present finding suggests that the codon R213 of 11betaHSD2 is a hot spot for mutations in this gene, as shown by the occurrence of an R213C point-mutation in several families unrelated to each other.

Kinetoplast DNA from Trypanosoma rangeli contains two distinct classes of minicircles with different size and molecular organization.

Trypanosomatids are characterized by the presence of kinetoplast DNA (kDNA), a peculiar form of mitochondrial DNA that consists of several thousand minicircles and a few dozen maxicircles catenated in a network. Within a species, the minicircles are known to differ in nucleotide sequence, but are homogeneous in size and always cross-hybridize. In all species of trypanosomatids, kDNA minicircles have at least one copy of a conserved 100-200 nucleotide region containing an almost invariant 'universal' 12-mer sequence (5'-GGGGTTGGTGTA-3'). We here report that Trypanosoma rangeli, a non-pathogenic parasite of man, contains two distinct classes of kDNA, minicircles called KP1 and KP2, which differ in size and molecular organization. Both were cloned and sequenced in both directions. KP2 was 1587 bases along and contained two copies of the conserved region as direct repeats 180 degrees apart. In contrast, KP1 had 1764 bases and showed a single conserved region. Moreover, KP1 differed further from KP2 and from most other previously sequenced trypanosomatid minicircles by containing a nucleotide substitution (5'-GGGGTTAGTGTA-3') in the 12-mer universal sequence tag. Polymerase chain reaction and hybridization studies suggest that the sequence of KP1 is very conserved in several other T. rangeli strains from Honduras, Colombia and Venezuela. It thus could provide a good target for the molecular diagnosis of infection with this parasite.

Hybridization screening of very short PCR products for paleoepidemiological studies of Chagas' disease.

Single strands of very short PCR products can be covalently immobilized to a slide and then easily detected by probe hybridization. In this work, the PCR product was a 70-nucleotide segment of ancient DNA, representing a portion of repeat mini-circle DNA from the kinetoplast of Trypanosoma cruzi, the infectious agent of Chagas' disease (American Trypanosomiasis). The target segment was initially established to be present in soft tissue samples taken from four "naturally" mummified Andean bodies using PCR followed by cloning and sequencing. Hybridization screening of the covalently immobilized PCR products positively identified products from 25 of 27 specimens of different tissues from these four mummies. The method appears to be ideal for the purpose of screening a large number of specimens when the target PCR product is very short.

DNA fingerprinting reveals relationships between strains of Trypanosoma rangeli and Trypanosoma cruzi.

Very little is known about the structure and sequence of the genomic DNA and kDNA of T. rangeli and no highly polymorphic markers are known. In this paper, we show that the Jeffreys' multilocal probe 33.15 produces characteristic DNA fingerprints with these trypanosomes. The multiband patterns can be used to differentiate T. cruzi from T. rangeli and for recognizing relationships between strains of the latter from widely different geographic areas and different hosts. The topology of a UPGMA phenetic tree constructed from band-sharing data suggests the existence of two groups of T. rangeli: one encompassing parasites from Central America and the northern part of South America and another with the parasites from southern Brazil. This splitting was confirmed by the use of both nuclear and kinetoplast unique sequence probes. Among strains of T. rangeli, band sharing was generally negatively correlated with geographical distance. This work confirms the usefulness of DNA fingerprints as a potent technique for the analysis of relationships in trypanosomatid populations.

Comparative effects of aspirin and enteric-coated aspirin on loss of chromium 51-labeled erythrocytes from the gastrointestinal tract.

Sodium chromate Cr 51 was used to label red blood cells of 19 healthy male volunteers, whose stools were collected for four days before and four days during oral administration of either uncoated (N = 9) or enteric-coated (N = 10) aspirin. Each subject received 2.925 gm/day of aspirin, in three equal doses separated by eight-hour intervals, for a total of seven days. During drug use, stools were collected on days 4 through 7. Fecal blood content, estimated by measuring radioactivity in the stools, was significantly higher (P less than 0.001) during use of either type of aspirin than at baseline, but losses measured during use of the coated aspirin (mean, 1.54 ml/day) were significantly lower (P less than 0.001) than those measured during use of the uncoated aspirin (mean, 4.33 ml/day). The two types of aspirin produced equivalent serum concentrations of salicylates. We conclude that enteric-coated aspirin reduces gastrointestinal blood loss.

Functional sexual dimorphism of the nucleolar organizer regions in the tuberomamillary nucleus.

The tuberomamillary nucleus (TM) is a cluster of magnocellular neurons that are located close to the mamillary recess or basal surface of the mamillary body (MBs) and is the only known source of histamine (HA). The nucleolar organizer regions (NORs) of the nervous cells of the subnuclei E1, E2 and E3 of the TM are quantified in this paper to discern the existence of possible sexual differences in the overall neuronal protein synthesis between male and female rats in two phases of the oestrous cycle (oestrous-dioestrous). We have used the argentic impregnation technique of the NORs (AgNORs) in both bilateral nuclei of the TM and found functional bilateral symmetry in its AgNORs. Furthermore, we demonstrate the existence of significant differences (P < 0.001) between sexes and between two phases of the oestrous cycle studied (oestrous-dioestrous) in the parameter of number of AgNORs per neuron. Hormonal manipulation of the TM in the rat during the two periods, postnatal or adult, shows the importance of the activational effect of the hormones on the activity of cellular protein synthesis.

Species specific detection of Trypanosoma cruzi and Trypanosoma rangeli in vector and mammalian hosts by polymerase chain reaction amplification of kinetoplast minicircle DNA.

Several groups have recently developed molecular tests for the detection of Trypanosoma cruzi, the causative agent of Chagas' disease. Polymerase chain reaction (PCR) amplification of kinetoplast minicircle DNA sequences appears to be the most sensitive method. However, the specificity of PCR-based diagnostic methods was challenged when the complete sequence of Trypanosoma rangeli DNA minicircles was discovered. In the present study, we conducted. PCR experiments using the S35/S36 primers in Rhodnius prolixus and Balb/c mice with single and mixed infections of T. cruzi and/or T. rangeli. In single infections, the profile of each trypanosome was easily distinguishable in haemolymph, salivary gland and intestinal tissues and faeces of insect vectors. In mixed infections of anterior intestine (where T. rangeli is more predominant than T. cruzi), the DNA amplification profile of both parasites was observed simultaneously. Conversely, only the T. cruzi profile was observed in rectal ampulla (where T. cruzi is more abundant than T. rangeli). In mice with single infections of T. cruzi or T. rangeli, the profiles of amplified DNA were easily distinguishable in each case. The T. cruzi profile was dominant in most mixed infections, probably due to the fact that T. cruzi minicircles are more abundant and consequently compete more eagerly for annealing with the S35/S36 primers. In cases of mixed infections where T. rangeli was initially more abundant than T. cruzi, the specific T. rangeli 760 bp band was present for 7 days after infection and then this band and others ranging from 300 to 450 bp disappeared and only the typical T. cruzi 330 bp band remained. The S35/S36 primers used in polyacrylamide gel electrophoresis (PAGE) detected T. cruzi specifically, and prevented misdiagnosis due to the presence of T. rangeli. This technique can also be used to identify parasites in different stages of the infection (acute or chronic) in vertebrate hosts and to localize the parasites in the insect vector.

The affinity of the lectins Ricinus communis and Glycine maxima to carbohydrates on the cell surface of various forms of Trypanosoma cruzi and Trypanosoma rangeli, and the application of these lectins for the identification of T. cruzi in the feces of Rhodnius prolixus.

Flagellates of Trypanosoma cruzi (stock Molino 1), obtained from the intestine of experimentally infected Rhodnius prolixus, grown in cellular or acellular culture, as well as from the blood of infected mice, were examined by a direct fluorescence test using the lectins RCA (Ricinus communis-120) and SBA (soy bean agglutinin; Glycine maxima), conjugated with fluorescein isothiocyanate, for the detection of beta-D-galactose and alpha,beta-N-acetyl-D-galactosamine on the membranes of the flagellates. The same reactions were carried out using Trypanosoma rangeli (stock San Agustin), obtained from the intestine, hemo-lymph or salivary glands of experimentally infected R. prolixus, as well as from cultures and from the blood of experimentally infected CFW mice. The results indicate that the membrane of T. rangeli in the salivary glands of the vector contains beta-D-galactose, but that this sugar is absent from all other developmental stages of this trypanosome. All stages of intestinal and cultured. T. cruzi presented positive reactions with RCA-FITC and SBA-FITC. The high specificity of this technique makes it useful for the examination of R. prolixus, previously used in xenodiagnosis of Chagas' disease and for the examination of intradomiciliary or sylvatic vectors in epidemiological surveys in areas where T. cruzi and T. rangeli coexist. Formaldehyde fixed samples can be examined months later and false reports due to T. rangeli can be avoided.

Neuroprotective effect of melatonin on brain damage induced by acute global cerebral ischemia in cats.

BACKGROUND: Melatonin has been proposed as a neuroprotective agent on the basis of its ability to function as a free radical scavenger, provided that lipoperoxidation and other free radical damage induced by reactive oxygen species resulting from cerebral ischemia are relevant pathophysiologic processes of ischemic neuronal damage. METHODS: The effects of melatonin or vehicle on neurologic deficit scores (Todd scale; maximal deficit score = 100, daily during 7 days after the ischemic episode) and neuronal population of hippocampal CA1-CA4 fields (cresyl violet stain, at the eighth day after the ischemic episode) were evaluated in adult male cats subjected to a 15-min period of acute global cerebral ischemia induced by cardiorespiratory arrest, and in cats subjected to a sham procedure. Continuous intravenous (iv) administration of either melatonin 10 mg/kg/h in 10% ethanol in saline or the vehicle alone (3 mL/kg/h) for 6 h starting 30 min after the end of the period of global cerebral ischemia was used as treatment. RESULTS: Global cerebral ischemia resulted in a severe loss of neurons in hippocampal CA1-CA4 fields (9, 13, 30 and 28% remaining neurons, respectively) of ischemic vehicle-treated cats in comparison with sham cats (100%). By contrast, remaining neurons in these regions were between 81 and 100% in the ischemic melatonin-treated cats, values that are nonsignificantly different as compared with sham cats. Values of remaining neurons in CA1-CA4 fields in ischemic melatonin-treated cats were significantly higher than those in ischemic vehicle-treated cats. Neurologic deficit scores in ischemic vehicle-treated cats (42-77 at day 1, 6-39 at day 7) were significantly higher than those in ischemic melatonin-treated cats (16-38 at day 1, 0-6 at day 7) on the days after the ischemic episode. CONCLUSIONS: Overall, the results support the neuroprotective effect of melatonin against the neuronal cerebral damage induced by acute global cerebral ischemia.

Unusual periodontal findings in an AIDS patient with Burkitt's lymphoma. A case report.

This report deals with a case of AIDS-related Burkitt's lymphoma in which some of the earliest findings were dental and periodontal in nature. The patient presented initially with painless and extremely loose teeth accompanied by progressive paresthesia of the lower jaw. Unique radiographic findings included extensive periodontal ligament space widening and bulbous, granuloma-like lesions about the apices of the teeth. These findings were associated with progressive tumor infiltration of the mandible and do not appear to be related to other reports of aggressive periodontitis associated with impaired immunologic functions in AIDS patients.