Pathophysiology of spontaneous coronary artery dissection: hematoma, not thrombus.

Spontaneous coronary artery dissection (SCAD) accounts for 1.7%-4% of all acute coronary syndrome presentations, particularly among young women with an emerging awareness of its importance. The demarcation of acute SCAD from coronary atherothrombosis and the proper therapeutic approach still represents a major clinical challenge. Certain arteriopathies and triggers are related to SCAD, with high variability in their prevalence, and often, the cause remains unknown. The objective of this review is to provide contemporary knowledge of the pathophysiology of SCAD and possible therapeutic solutions.

Quantitative and Qualitative Characteristics of Atherosclerotic Plaques on Carotid Arteries in Patients with Antiphospholipid Syndrome: The Role of MDCT Angiography.

INTRODUCTION: Antiphospholipid syndrome (APS) is an autoimmune disease characterised by arterious and venous thrombosis, miscarriage, and the presence of antiphospholipid antibodies (aPL) in the blood. As we know, APS is also characterised by accelerated atherosclerotic degeneration with an increased risk of thrombosis in all blood vessels, including the carotid arteries. Carotid artery stenosis can manifest in many different ways. The aim of this study is to present the results of our multidetector computerised tomography angiography (MDCTA) analysis of the carotid arteries in patients with primary and secondary APS compared with a control group. MATERIALS AND METHODS: This study examined 50 patients with primary antiphospholipid syndrome (PAPS) and 50 patients with secondary antiphospholipid syndrome (SAPS). The results were compared with a control group also comprising 50 patients. The groups were analysed with respect to age, sex and the presence of well-established risk factors for vascular disease. The study was conducted using MDCTA, where we analysed the quantitative and qualitative (morphologic) characteristics of carotid artery lesions. RESULTS: Patients from the control group had significantly elevated levels of cholesterol and triglycerides in comparison with patients with PAPS and SAPS (p < 0.001 and p < 0.05). The results show that carotid artery lesions were significantly more common in patients with APS (PAPS, n = 40, CI95: 0.50-0.75, p = 0.0322 and SAFS, n = 54, CI95: 0.59-0.80, p = 0.0004) than within the control group (n = 23). There was a statistically significant difference between patients with APS and the control group with respect to lesions in the distal segments (n = 27, CI95: 0.67-0.95, p = 0.0001), bulbi and proximal segments (n = 21, CI95: 0.84-1.00, p = 0.000005). The number of patients with one lesion (L) (n = 27) was significantly greater than the number of those with three (n = 10, CI95: 0.56-0.86, p = 0.0051) or four (n = 3, CI95: 0.73-0.98, p = 0.00001) lesions. There were also more patients with two lesions (n = 24) than those with four (n = 3) (CI95: 0.71-0.97, p = 0.00005). Carotid artery stenosis was shown as a percentage of the carotid artery lumen diameter (%DS). Stenosis of up to 30%, was more common in patients in the PAPS group (n = 12) than in the control group (n = 3) (CI95: 0.52-0.96, p = 0.0201), while the SAPS group (n = 17) had an even larger disparity (CI95: 0.62-0.97, p = 0.0017). We observed a highly significant difference in the frequency of stenoses between 30% and 50% DS between the PAPS group (n = 24) and the control group (n = 7) (CI95: 0.59-0.90, p = 0.0023), as well as the SAPS group (n = 30) (CI95: 0.65-0.92, p = 0.0002). A qualitative analysis of plaque morphology revealed that patients with PAPS had significantly more soft tissue lesions (n = 23) compared with calcified lesions (n = 2) (CI95: 0.74-0.99, p = 0.00003), as well as more mixed plaques (n = 9) and calcified plaques (n = 2) (CI95: 0.48-0.98, p = 0.0348). Patients within the SAPS group had significantly more soft tissue (n = 35) than calcified lesions (n = 3) (CI95: 0.79-0.98, p = 0.00000021), as well as more mixed lesions (n = 21) compared with calcified (n = 3) (CI95: 0.68-0.97, p = 0.0002). CONCLUSIONS: Our study shows that subclinical manifestations of carotid artery lesions were more common in patients with APS. We came to the conclusion that MDCTA is an accurate diagnostic method because it is a safe method that provides us with a great quantity of accurate information about the characteristics of atheromatous plaques, which aids us in the further planning of treatment for patients with APS.

Morphometric and volumetric analysis of lacrimal glands in patients with thyroid eye disease.

Assessment of morphometric and volumetric changes in lacrimal glands in thyroid eye disease, its clinical manifestations in relation of disease progression. Retrospective volumetric analysis included both genders and was performed on total of 183 patients - 91 patients with diagnosed Grave's disease and thyroid eye disease and 92 patients without Grave's disease and thyroid eye disease who underwent multidetector computed tomography (MDCT) examination in routine daily work according to other medical indications. In the group of females, there was statistical significance between patients with thyroid eye disease and controls who were smoking and had body weight gain. We found statistical significance in volumetric enlargements for both orbits in both genders for the patients group when compared to controls. There was also statistical significance in morphometric characteristics for the lacrimal gland diameters measured. Determination planimetric morphometric parameters of importance were coronary height of lacrimal gland of the right eye, coronary height of lacrimal gland of the left eye and coronary width of lacrimal gland of the left eye for the group of males. In a group of females the established determination parameters of importance were the coronary height of lacrimal gland of the left eye, the axial width of lacrimal gland of the left eye, volume of lacrimal gland of the right eye and the volume of lacrimal gland of the left eye. When we compared the displaced lacrimal gland coming forward (proptosis) in time progressing disease between group of patients and controls, we also found statistical significant connection. Evaluation of lacrimal gland volumetric and morphometric data may increase validity of defining this anatomical substrate and its morphology disruption as liable tool for thyroid eye disease progression follow up and treatment planning and outcome.

The long-term effects of maternal deprivation on the number and size of inhibitory interneurons in the rat amygdala and nucleus accumbens.

Introduction: There is an increasing evidence supporting the hypothesis that traumatic experiences during early developmental periods might be associated with psychopathology later in life. Maternal deprivation (MD) in rodents has been proposed as an animal model for certain aspects of neuropsychiatric disorders. Methods: To determine whether early-life stress leads to changes in GABAergic, inhibitory interneurons in the limbic system structures, specifically the amygdala and nucleus accumbens, 9-day-old Wistar rats were exposed to a 24 h MD. On postnatal day 60 (P60), the rats were sacrificed for morphometric analysis and their brains were compared to the control group. Results: Results show that MD affect GABAergic interneurons, leading to the decrease in density and size of the calcium-binding proteins parvalbumin-, calbindin-, and calretinin-expressing interneurons in the amygdala and nucleus accumbens. Discussion: This study indicates that early stress in life leads to changes in the number and morphology of the GABAergic, inhibitory interneurons in the amygdala and nucleus accumbens, most probably due to the loss of neurons during postnatal development and it further contributes to understanding the effects of maternal deprivation on brain development.

The role of cadmium in the pathogenesis of myeloid leukemia in individuals with anemia, deficiencies in vitamin D, zinc, and low calcium dietary intake.

Iron deficiency, vitamin D deficiency and low calcium diet are frequent health problems with severe long- term consequences. Upon absorption from the duodenum, cadmium binds to transferrin, and cells with the highest density of transferrin receptor 1 (TfR1) take up the majority of the circulating cadmium. Nowadays, it is clear that individuals with iron deficiency anemia have increased blood levels of cadmium because of higher absorption rate, mediated by divalent metal transporter 1 (DMT1). However, the transient receptor potential vanilloid receptor 6 (TRPV6), known as a calcium carrier, is able to bind and transport cadmium as well. In the case of low calcium diet or vitamin D deficiency, TRPV6 may be overexpressed in the intestine and kidney tubules and absorbs (re-uptake in the case of renal tubules) cadmium in larger quantities, resulting in an increased cadmium blood levels. We speculate that the final event in the case of low calcium dietary diet and/or vitamin D deficiency is similar to what is observed in the case of iron deficiency, that cells with the highest levels of TfR1 (for example, megakaryocyte/erythrocyte progenitors and pro-erythroblasts) take up most of the circulating cadmium, which is powerful malignancy inductor, leading to appearance of acute myeloid leukemia (AML).

Medullary branches of the vertebral artery: microsurgical anatomy and clinical significance.

BACKGROUND: Since the medullary arteries are of a great neurologic and neurosurgical significance, the aim was to perform a detailed microanatomic study of these vessels, as well as of the medullary infarctions in a group of patients. METHODS: The arteries of 26 halves of the brain stem were injected with India ink and gelatin, microdissected and measured with an ocular micrometer. Neurologic and magnetic resonance imaging (MRI) examinations were performed in 11 patients. RESULTS: The perforating medullary arteries, averaging 6.7 in number and 0.26 mm in diameter, most often originated from the anterior spinal artery (ASA), and rarely from the vertebral (VA) (38.5%) and the basilar artery (BA) (11.6%). They supplied the medial medullary region. The anterolateral arteries, 4.8 in number and 0.2 mm in size, most often arose from the ASA and PerfAs, and nourished the anterolateral region. The lateral arteries, 2.2 in number and 0.31 mm in diameter, usually originated from the VA and the posterior inferior cerebellar artery (PICA). They supplied the lateral medullary region. The dorsal arteries, which mainly arose from the PICA and the posterior spinal artery (PSA), nourished the dorsal region, including the roof of the 4th ventricle. The anastomotic channels, averaging 0.3 mm in size, were noted in 42.3%. Among the medullary infarctions, the lateral ones were most frequently present (72.8%). CONCLUSION: The obtained anatomic data, which can explain the medullary infarctions symptomatology, are also important in order to avoid damage to the medullary arteries during neurosurgical and neuroradiologic interventions.

Effects of Maternal Deprivation on the Prefrontal Cortex of Male Rats: Cellular, Neurochemical, and Behavioral Outcomes.

Stressful events experienced during early life are associated with increased vulnerability of developing psychopathology in adulthood. In the present study, we exposed 9-day-old Wistar rats to 24 h maternal deprivation (MD) with the aim to investigate the impact of early life stress (ELS) on morphological, biochemical, and functional aspects of the prefrontal cortex (PFC), a brain region particularly sensitive to stress. We found that in the superficial medial orbital cortex (MO), young adult male rats had reduced density of GAD67 and CCK immunopositive cells, while the rostral part of the ventral lateral orbital cortex (roVLO) showed a decrease in the density of GAD67 immunopositive cells in both superficial and deep layers. In addition, the superficial rostral part of area 1 of the cingulate cortex (roCg1) and deep prelimbic cortex (PrL) was also affected by MD indicated by the reduction in PV immunopositive cellular density. Furthermore, MD induced upregulation of brain-derived neurotrophic factor (BDNF), while it did not affect the overall expression of Iba1 in neonatal or young adult PFC as measured by Western blot, however, microglial activation in young adult MD rats was detected immunohistochemically in deep layers of MO and infralimbic cortex (IL). Interestingly, when young adult male rats were subjected to a behavioral flexibility test in a T-maze, MD rats showed a subtle impairment in T-maze reversal learning indicating a mildly affected PFC function. Taken together, our findings demonstrated that MD reduced the density of interneurons and induced microglial activation, in particular, PFC areas at young adulthood, and could alter synaptic plasticity accompanied by PFC dysfunction.

Glucocorticoid levels after exposure to predator odor and chronic psychosocial stress with dexamethasone application in rats.

This study was conducted to explore the effects of specific psychosocial paradigm on predator animal posttraumatic stress model and to test the hypothesis that psychosocially stressed rats would exibit abnormal levels of cortisol and a larger suppression of cortisol levels after the application of dexamethasone. Animals were divided in two groups: experimental and control groups. The experimental group was exposed to two types of stressors: acute immobilization stress, and combined predator stress and daily social stress with application of dexamethasone. Blood sampling was performed at three different times. We found statistically significant results after analyzing the differences between cortisol levels in different times of blood sampling in the group of animals exposed to stress with dexamethasone application. Statistical significance was found when we compared the experimental group with the control group in terms of elevated cortisol levels during blood sampling after stress paradigm exposition. Many significant disruptions in the functioning of the hypothalamic-pituitary-adrenal axis were observed, such as decrease in basal cortisol levels and enhanced dexamethasone-induced inhibition of cortisol levels. These findings are important because their impact can translate to human individuals with posttraumatic stress disorder, which is the most important role of every animal model in research.

Clinical significance of blood supply to the internal capsule and basal ganglia.

Although the general vascular supply of the basal ganglia and internal capsule is well known, precise data are lacking regarding the variations of the vascular territories in the two regions. Twelve hemispheres were studied following an injection of coloured ink into the main cerebral arteries, namely the anterior cerebral (ACA), middle cerebral (MCA), anterior choroidal (AChA) and posterior cerebral artery (PCA). Serial sections of the injected hemispheres were taken in the axial or coronal plane. In 75% of the hemispheres, ACA perforators were seen to supply the inferomedial part of the head of the caudate nucleus and the anterior limb of the internal capsule, as well as the anterior and inferior portions of the putamen and globus pallidus. The MCA vessels perfused the superolateral part of the head and body of the caudate nucleus, the superior part of the entire internal capsule, most of the putamen and part of the globus pallidus. The AChA perforators perfused the medial segment of the globus pallidus, the inferior part of the posterior limb, the retrolenticular and sublenticular portions of the internal capsule, and occasionally its genu. The same segment of the globus pallidus and the inferior part of the genu of the internal capsule were most likely supplied by the perforators of the internal carotid artery. A predominance of ACA territory was noticed in one specimen (8.33%) and a predominance of MCA territory in two specimens (16.67%). The obtained anatomical data may help radiologic determination of perforators involved in ischemic events, as well as a better understanding of the neurological deficits in the same events.

Common features of the cerebral perforating arteries and their clinical significance.

BACKGROUND: The perforating vessels supply very important regions of the brain stem and diencephalon, as well as the basal ganglia and internal capsule. Some of their micro-anatomical characteristics are still not well known. The aim of this study was to examine and evaluate the features of all the perforating vessels. METHODS: The arteries of 24-32 cerebral hemispheres, diencephalons and halves of the brain stem were injected with India ink mixture or methylmethacrylate, and microdissection was performed or the vascular casts were produced and examined under the sterescopic microscope. RESULTS: It was noticed that the perforators ranged from 0 to 14 in number, with the smallest mean value (1.1) for the diencephalic perforators and the largest one (8.1) for the lenticulostriate arteries. The smallest mean diameter (175 µm) was found in the group of the perforators of the anterior communicating artery, whereas the largest one is related to the Heubner's artery (668 µm), the diencephalic thalamoperforating vessels (562 µm), the premamillary vessel (489 µm) and the lenticulostriate arteries (469 µm). The perforators most frequently originated from the pial branches of the basilar artery (91.7 %) and of the posterior cerebral artery (59.4 %). The common stems were most often formed by the perforators of the basilar (79.2 %), posterior cerebral (75.0 %) and middle cerebral arteries (40.6 %). Some perforators arose close to or from the terminal divisions, the branching sites or the junctions of the parent arteries, where the saccular aneurysms most often develop. The anastomoses among the perforators were present in a range from 6.3 % to 53.2 %. CONCLUSIONS: The micro-anatomical data obtained may be useful for neurosurgeons when operating at the base of the brain, as well as for a neurological and radiological evaluation of the perforators in the occlusive cerebrovascular disease, or in the cases of an aneurysm, arteriovenous malformation (AVM) or tumour presence.

The influence of sex hormones on anterior cruciate ligament ruptures in males.

PURPOSE: The purpose of this study is to determine the difference in the concentrations of testosterone, 17-ß estradiol and progesterone between male patients with and without ACL rupture, as well as the possible effect of these hormones on generalized joint laxity. METHODS: Male subjects with non-contact knee joint injury were included in this study. Two groups were formed: the examined group, consisting of subjects with ACL rupture and the control group consisting of patients without ACL rupture. After this, the patients from these two groups were paired off on the basis of three factors, level of professional involvement in sports (including the type of sports activity), left or right side of the body and the age of the subjects. In the end, there were 29 pairs (58 subjects). The concentration of sex hormones was determined from saliva specimens with the aid of the Salimetrics enzyme immunoassay. The testing of generalized joint laxity was performed with the aid of the "laxity score" according to Beighton et al. RESULTS: Subjects with ACL rupture have highly statistically significantly greater concentrations of testosterone (p < 0.01), statistically significantly greater concentrations of 17-ß estradiol (p < 0.05), and a highly statistically significantly greater generalized joint laxity score than subjects with an intact ACL (p < 0.01). CONCLUSION: Increased concentrations of testosterone or 17-ß estradiol may be a risk factor leading to ACL rupture. Also, generalized joint laxity may be a factor leading to ACL rupture, but none of the monitored hormones can be set down as the cause of its existence. Young male athletes with higher concentrations of testosterone and greater hyperelasticity should plan preventive programs of physiotherapy for ACL preservation since they present a vulnerable group susceptible to ACL rupture. LEVEL OF EVIDENCE: Diagnostic study, Level II.

The influence of sex hormones on anterior cruciate ligament rupture: female study.

PURPOSE: The purpose of this study was to determine the difference in the concentrations of testosterone, 17-ß estradiol and progesterone between female patients with and without ACL rupture and the possible effect of these hormones on generalised joint laxity. METHODS: Female subjects with non-contact knee joint injury were included in this study. They were divided into two groups: the examined group, consisting of female subjects with ACL rupture, and the control group, consisting of female patients without ACL rupture. In the next step, the patients from these two groups were paired off on the basis of three factors: the level of professional sports involvement (including the type of sports activity), the side of the body where the injury had occurred (left or right) and the age of the subjects. In the end, there were 12 pairs (24 subjects). The concentrations of sex hormones were established from saliva specimens with the aid of the Salimetrics enzyme immunoassay. Generalised joint laxity was tested with the aid of the "laxity score" according to Beighton, Solomon and Soskolne. RESULTS: Female subjects with ACL rupture had significantly lower concentrations of testosterone (p < 0.01), significantly lower concentrations of 17-ß estradiol (p < 0.05) and significantly lower concentrations of progesterone (p < 0.01) than female subjects with intact ACL. CONCLUSIONS: Decreased concentrations of testosterone, 17-ß estradiol or progesterone may be a risk factor leading to ACL rupture. The concentrations of these hormones do not affect generalised joint laxity. Additional research on a larger group of patients is necessary to further determine the effects of these hormones on generalised joint laxity and ACL ruptures. Young female athletes with lower concentrations of sex hormones are more prone to anterior cruciate ligament rupture which is why they need to reduce their sports activities during the pre-ovulatory phase of the menstrual cycle, when these concentrations are additionally reduced.

[The role of the posterior tibial slope on rupture of the anterior cruciate ligament].

BACKGROUND/AIM: Posterior tibial slope is one of the most citated factors wich cause rupture of the anterior cruciate ligament (ACL). The aim of this study was to determine the association of a greather posterior tibial slope on the lateral condyle, that is a lesser posterior tibial slope on the medial condyle, with ACL rupture. METHODS: The patients were divided into two groups. The study group included the patients with chronic instability of the knee besause of a previous rupture of ACL. The control group included the patients with knee lesion, but without ACL rupture. Posterior tibial slope measuring was performed by sagittal MR slices supported by lateral radiograph of the knee. We measured posterior tibial slope on lateral and medial condyles of the tibia. Using these values we calculated an average posterior tibial slope as well as the difference between slopes on lateral and medial condyles. RESULTS: Patients with ACL rupture have highly statistically significantly greather posterior tibial slope (p < 0.01) on lateral tibial condyle (7.1 degrees : 4.5 degrees) as well as statistically significantly lesser posterior tibial slope (p < 0.05) on medial tibial condyle (5.9 degrees : 6.6 degrees) than patients with intact ACL. CONCLUSION: Great posterior tibial slope on lateral tibial condyle associated with the small posterior tibial slope on the medial tibial condyle, that is a positive differentce between lateral and medial tibial condyles are factors wich may cause ACL rupture.

Morphometric analysis, region of supply and microanatomy of the lenticulostriate arteries and their clinical significance.

The lenticulostriate arteries (LSA) and their microanatomy, region of supply and atherosclerosis were examined in 24 microdissected brains, arterial casts, and histological specimens. The LSA ranged from 2 to 12 in number and from 0.10 mm to 1.28 mm in diameter. They always arose from the initial segment of the middle cerebral artery (MCA), often from the MCA leptomeningeal branches (38.24%), and rarely from the insular segment (2.94%). They always originated as individual branches, often (61.76%) with their own common stems. In two hemispheres we found that the LSA supplied either a larger or a smaller portion of the basal ganglia and internal capsule than usual. The number of twigs to the innominate substance (substantia innominata) (3-11), and their diameters (0.07-0.30 mm), has been described for the first time, to our knowledge. Microatheromas were found in two LSA. Data about the LSA microanatomy and territory could form the basis of safer neurosurgery, more accurate neuroimaging evaluation, and precise neurological diagnosis in patients with focal ischemic lesions in the basal ganglia and internal capsule.

[Congenital interruption of the inferior vena cava: a case report].

INTRODUCTION: Congenital interruption of the inferior vena cava is a rare entity. It can be associated with other organ and system malformations, especially the cardiovascular system. Usually, patients are asymptomatic and the anomaly is therefore diagnosed incidentally. In some cases it can be of clinical importance. CASE OUTLINE: A 22-year-old female was diagnosed during evaluation of cardiac symptoms. Clinical examination showed normal findings. CT angiography and cavography were used to establish the diagnosis of inferior vena cava interruption. Since there was no need for surgical intervention the patient was discharged in good condition. During a five year follow-up no significant health problems were noted with patient leading normal life. CONCLUSION: Diagnostic tools used to establish the diagnosis of inferior vena cava interruption are ultrasonography, as well as CT and MR angiography and phlebography. Mediastinal masses found on chest roentgenogram must raise suspicion of inferior vena cava interruption. Although the anomaly is rare, physicians should be aware of the infrahepatic interruption of the IVC, especially because it can present at any age.

Microanatomy of the intrachoroidal vasculature of the lateral ventricle.

OBJECTIVE: Intraventricular surgery requires a detailed knowledge of the microanatomy of the choroid plexus vasculature. METHODS: Twenty choroid plexuses were microdissected, and two additional plexuses were prepared for microscopic examination. RESULTS: The choroid plexus was perfused primarily by the anterior choroidal artery (AChA) and the lateral posterior choroidal artery (LPChA). The AChA, which averaged 650 microm in diameter, most often (in 75% of cases) divided into the medial and lateral trunks, which averaged 450 microm in diameter. The medial trunk gave off the bush-like intrachoroidal branches, whereas the lateral trunk divided into the parallel arteries. The inferior LPChA was present in 50% of the hemispheres, both the inferior and superior LPChAs in 40%, and their common trunk in 10%. In 40%, the LPChA, which averaged 670 microm in diameter, divided into the terminal trunks, with a mean diameter of 490 microm. The anastomoses involving the trunks of the LPChA and other choroidal arteries averaged 310 microm in diameter. All primary intrachoroidal branches of the AChA and LPChA were divided into three groups. The parallel branches, which averaged from 220 to 230 microm in diameter, coursed along the lateral part of the choroid plexus. The tortuous glomus vessels, which averaged 310 microm in size, originated from the AChA (45%), the LPChA (15%), or both (40%). The bush-like vessels, with a mean diameter between 155 and 190 microm, ramified into smaller twigs, up to the intrachoroidal capillaries. CONCLUSION: The data obtained on the microanatomy of the intrachoroidal vasculature may have certain neurosurgical implications.

Microsurgical anatomy of the perforating branches of the vertebral artery.

BACKGROUND: There is limited data in the literature related to the microanatomic features of the perforating branches of the vertebral artery. METHODS: The 44 vertebral arteries and their branches were injected with india ink or a radiopaque substance and examined under the stereoscopic microscope. RESULTS: The perforating arteries were noted to range in number from 1 to 11 (mean, 6.5) and in diameter between 100 microm and 520 microm (average, 243 microm). They arose from the vertebral artery (VA) (54.54%), 8 from the right, the left or both VAs. The anterior spinal artery (ASA), which was singular (81.82%), duplicated (13.64%), or plexiform (4.55%), always gave rise to the perforators. The vascular roots of the ASA were the source of the perforators in 95.45% of the brains. The latter vessels arose from the anterolateral arteries in 50% of the cases. The anastomoses involving the perforators, which were present in 40.91% of the brains, varied in diameter between 100 microm and 350 microm (mean, 169 microm). The perforating vessels gave rise to the side branches in 95.45% of the brains that varied in diameter from 100 microm to 300 microm (average, 161 microm). The perforators usually entered the foramen cecum and the anterior median sulcus, and then continued close and parallel to the raphe of the medulla. The perforators can be compressed by a VA aneurysm, which was found in one among the 71 examined patients with cerebral aneurysms. CONCLUSIONS: The obtained data give additional information about the vascular anatomy of the pontomedullary region.