Sponge artifact in biopsy specimens

We describe a sponge-induced artifact in histologic sections of small biopsy specimens. The artifacts are angulated, often triangular holes within the tissue. They appear to be introduced as individual sponge barbs become embedded in the perimeter of biopsy specimens during tissue processing. The artifact is generally of little importance, but in certain specimens, such as needle biopsies of the kidney or liver, it may occasionally obscure important information. Other methods, such as lens paper wrapping, may be superior in these situations. The utility of the tissue cassette sponge, in most situations, outweighs the artifact.     

Early ultrastructural changes of antral mucosa with aspirin in the absence of Helicobacter pylori.

AIMS--To describe the ultrastructural changes that occur in human antral mucosa following direct application of aspirin in volunteers without Helicobacter pylori infection. METHODS--Ten healthy male volunteers without H pylori infection underwent three consecutive endoscopies (at zero, one and five hours). At the first endoscopy, two biopsy specimens were obtained (one for histology and the other for electron microscopy (EM)). At subsequent endoscopies, a single biopsy specimen was obtained for EM. A 50 ml solution of aspirin (concentration 3 mg/ml) was applied to the antral mucosa at the first endoscopy in five subjects; the other five subjects received 50 ml distilled water (placebo). RESULTS--The ultrastructural appearance of the first biopsy specimen in all subjects and subsequent biopsy specimens in the placebo treated subjects was normal. The aspirin treated group had evidence of intercellular oedema, widening of capillary fenestrae, rupturing of apical membranes, and dilatation of endoplasmic reticulum and mitochondria after one hour; these changes were more marked at five hours. Tight junctions were maintained. CONCLUSION--This is the first study to describe the early ultrastructural changes in antral mucosa induced by aspirin in subjects without H pylori infection.     

Physiological saline is not a biocompatible peritoneal dialysis solution

We have previously demonstrated that daily exposure to dialysis fluid results in significantly increased peritoneal lymphatic flow. In this study, we investigated if daily intraperitoneal infusion of saline (isotonic, glucose free) could cause similar changes. METHODS: Sixteen male SD rats received daily infusion (i.p.) of 20 ml saline for ten days (Saline group). Twenty-four hours after the last infusion, a 4 hour dwell study using 25 ml 3.86% glucose dialysis solution with frequent dialysate and blood sampling was done in each rat as well as in rats which did not receive daily infusion (Control, n=8). Radiolabeled human albumin (RISA) was added to the solution as an intraperitoneal volume marker. Radioactivity, glucose, urea, sodium, and potassium were measured for each sample. In a separate study, the RISA absorption to peritoneal tissue was also determined. RESULTS: The net ultrafiltration was significantly decreased in the daily infusion group (p<0.05). However, the apparent volume at 3 minutes of the dwell was markedly increased; this was due to a significant increase in the RISA binding (1.5-12.0% in the Saline group vs. 0.45-1.12% in the Control group) to peritoneal tissues as assessed by measurement of RISA recovery at 3 min of the dwell. This resulted in a significant overestimation both of the intraperitoneal volume (IPV) at 3 min and the (apparent) fluid absorption rate (as estimated by the transport of RISA out of peritoneal cavity): 0.087+/-0.026 ml/min in the Saline group vs. 0.052+/-0.007 ml/min in the Control group, p<0.001. The direct lymphatic flow as estimated by the clearance of RISA to plasma (which should not be affected by the RISA binding) also increased markedly (0.021+/-0.005 ml/min in the Saline group vs. 0.008+/-0.001 ml/min in the control group). There was no significant difference in the D/P values for small solutes (urea, sodium, potassium, urate) and D/D0 for glucose between the two groups. CONCLUSIONS: 1) Daily infusion of physiological saline into peritoneal cavity may increase the peritoneal lymphatic flow; 2) The significant (apparent) increase in IPV shortly after infusion may suggest increased RISA binding to peritoneal tissues (which may be related to the damage of the tissues, and results in overestimation of the peritoneal fluid absorption rate); 3) Saline is not a biocompatible peritoneal dialysis solution, and should therefore not be used as a control or flush solution.     

Postcapillary venule-like transformation of peritubular capillaries in acute renal allograft rejection. An ultrastructural study.

To describe ultrastructural changes in renal peritubular capillaries during acute allograft rejection, biopsy specimens from five allografts with acute rejection were studied electron microscopically and compared with control specimens. The thickness, cross-sectional area, and luminal circumference of capillary endothelium were estimated morphometrically. Thickening of endothelial cells, loss of fenestration, increase in endothelial cell organelles, increased adherence and passage of lymphocytes and monocytes, and defects in the endothelial lining were demonstrated. The luminal circumference representing the size of the capillary was not changed. A balloonlike fragmentation of endothelial cells was sometimes observed around lymphocytes, suggesting cytotoxic injury to capillaries. The observations indicate that during rejection-induced endothelial activation, peritubular capillaries exhibit postcapillary venule-like transformation that enhances the influx of inflammatory cells into the kidney allograft.     

Pulmonary capillary recruitment in response to hypoxia in healthy humans: A possible role for hypoxic pulmonary venoconstriction?

We examined mechanisms by which hypoxia may elicit pulmonary capillary recruitment in humans. On separate occasions, twenty-five healthy adults underwent exposure to intravenous saline infusion (30 ml/kg ～ 15 min) or 17-h normobaric hypoxia ( [FIO2 = 12.5%). Cardiac output (Q) and pulmonary capillary blood volume (Vc) were measured before and after saline infusion and hypoxic-exposure by a rebreathing method. Pulmonary artery systolic pressure (sPpa) and left ventricular (LV) diastolic function were assessed before and after hypoxic-exposure via echocardiography. Saline infusion increased Q and Vc (P < 0.05) with no change in Vc/Q (P = 0.97). Hypoxic-exposure increased Vc (P < 0.01) despite no change in Q (P = 0.25), increased sPpa (P < 0.01), and impaired LV relaxation. Multiple regression suggested that ～ 37% of the hypoxia-mediated increase in Vc was attributable to alterations in Q, sPpa and LV diastolic function. In conclusion, hypoxia-induced pulmonary capillary recruitment in humans is only partly accounted for by changes in Q, sPpa and LV diastolic function. We speculate that hypoxic pulmonary venoconstriction may play a role in such recruitment.     

Uttrastructure of skin biopsy specimens in lysosomal storage diseases: Common sources of error in diagnosis

Common sources of error in the diagnosis of lysosomal storage diseases by ultrastructural examination of skin specimens have been identified in a series of biopsies from 72 patients. Four principal factors have emerged as leading pitfalls and sources of error in diagnosis. First, the skin biopsy technique itself may lead to alterations of normal skin ultrastructure. Second, artifacts may be produced during fixation and preparation of tissue for electron microscopy. Third, cellular organelles and structures normally present in human skin may be mistakenly interpreted as pathological. Fourth, the use of cultured skin fibroblasts for ultrastructural identification of storage material is often accompanied by artifacts induced in tissue culture and is not recommended. Recognition of these common problems may aid interpretation of the fine structure of skin abnormalities. Furthermore, when skin biopsy specimens are used as the primary source of diagnostic material, correlation of both skin ultrastructure and assay for specific lysosomal enzymes in cultured dermal fibroblasts will facilitate diagnostic accuracy.     

Clinical, light, and electron microscopy findings in idiopathic haematuria

The electron microscopic findings are reported in detail in 20 patients submitted to renal biopsy with the major complaint or clinical finding of gross or microscopic haematuria. The lesions were classified histologically into four groups: group 1, minor glomerular alterations; group 2, focal mesangial thickening and/or cellular proliferation; group 3, diffuse mesangial proliferation; and group 4, other lesions. The major ultrastructural alterations included irregularity in thickness and density of the capillary basement membrane, with apparent discontinuity and bi- or multilaminar splitting of the lamina densa. There were varying degrees of foot process fusion, visceral epithelial polykaryocytosis, and granular deposits related to the capillary basement membrane. Densities were found in the mesangial basement membrane-like material, which was often markedly increased in quantity. A few microtubular aggregates were observed in endothelial cell cytoplasm. Changes consistent with acute diffuse proliferative and membrano-proliferative glomerulonephritis were also seen. The significant clinical findings, histological groups, and ultrastructural changes are correlated.     

The effect of alloxan, and alloxan-induced diabetes on the kidney

Alloxan is known to induce diabetic renal changes as well as causing nephrotoxic alterations. However, no ultrastructural study has been performed to differentiate diabetic verses toxic affects of alloxan to the tubule and/or glomerulus. Therefore the present study used the "protected" kidney model to prevent one kidney from being exposed to the alloxan while allowing the other to receive the drug immediately. In all experimental animals the right renal hilum was gently occluded for 5 minutes and then released. This was performed prior to the injection of alloxan. Subsequently, the left renal hilum was occluded at the time of, and for 5 minutes after, alloxan administration (40 mg/kg i.v.). The experimental rats were divided into three groups: untreated diabetics, diabetics treated with protamine-zinc-insulin, and alloxan-treated rats that failed to become diabetic. Three groups of controls were included: one group received an equal volume of saline diluent as the experimental rats but no clamping of either renal hilum; another group received the saline and had the left renal hilum occluded for 5 minutes; and a third group had both the right and left renal hila occluded. All animals were followed and sacrificed after 9 weeks. Endogenous creatinine clearance did not change among groups. Alloxan-treated nondiabetic rats displayed marked interstitial nephritis in unprotected kidneys, while protected kidneys were normal. The diabetic state resulted in mesangial proliferation and focal glomerular basement membrane thickening as well as glomerular capillary endothelial abnormalities and visceral epithelial foot-process fusion. The endothelial changes consisted of focal areas showing a reduction in the size of endothelial fenestrae. All glomerular changes were ameliorated by insulin treatment. We conclude: 1) alloxan per se is distinctly nephrotoxic; and 2) the glomerular endothelium and epithelium are involved early in the course of experimental diabetes.     

Artifactual hydropic degeneration in skin biopsy specimens immersed in saline: a light and electron microscopic study

An artifact, consisting of hydropic degeneration of the basal cells and subepidermal bulla formation, is described in skin-punch biopsy specimens immersed in normal saline. The histologic appearance and ultrastructural features of this artifact are discussed. Proper handling of skin specimens intended for immunofluorescent studies is stressed. Similarities are noted between the histologic and ultrastructural findings of this artifact and of epidermolysis bullosa simplex and pathologic hydropic degeneration, as is seen in lupus erythematosus.     

Ultrastructural abnormalities of liver cells in Reye's syndrome

Electron microscopic observations were made on liver biopsy specimens from nine infants and children diagnosed as having Reye's syndrome by clinical, laboratory, and light microscopic criteria. In addition to excessive fat content, mitochrondrial abnormalities were the most frequent abnormal finding in the liver. However, no correlation could be established between the severity of mitochondrial changes and clinical or biochemical data, and two patients with low levels of the first two urea cycle enzymes showed only mild mitochondrial abnormalities. Nonspecific or artifactual factors have been suggested to explain the mitochondrial changes. However, the data of this study suggest that most of the reported mitochondrial abnormalities are not artifacts, and that they can be helpful in the ultrastructural diagnosis of Reye's syndrome.     

Ultrastructural Changes Suggestive of Focal Segmental Glomerulosclerosis in Atypical Minimal Change Nephrotic Syndrome

In an attempt to recognize early stages of focal segmental glomerulosclerosis (FSGS) in patients with a clinical course suggesting a diagnosis other than minimal change disease (MCD) and normal histology, or minor, nondiagnostic changes on light microscopy (LM), we used a protocol for systematic and extensive electron microscopy (EM) examination of kidney biopsies obtained from such patients. By this method ultrastructural pathology was found in 8 patients. These changes were localized, involving only portions of single glomerular segments. The findings included mild to moderate increase of the mesangial matrix, focal wrinkling of the capillary basement membrane, and early obliteration of the normal architecture of individual capillary loops, as well as electron-dense deposits in a mesangial and subendothelial distribution. Of these 8 patients, 2 are at present in remission without therapy (in 1, following therapy with cyclophosphamide); 3 are in remission on steroid therapy; 1 developed massive proteinuria during pregnancy, after a spontaneous remission lasting almost 2 years; 1 patient advanced to terminal renal failure 3 1/2 years after biopsy; and 1 died of sepsis 1 month after biopsy. We believe that the ultrastructural changes found may represent early or mild FSGS and that the protocol described can add valuable information in clinically worrisome patients in whom renal histology appears normal.     

Tubuloreticular inclusions in systemic lupus pneumonitis. Report of a case and review of the literature

Tubuloreticular inclusions have been described with sufficient frequency in certain tissues of patients with systemic lupus erythematosus (SLE) to make the finding of such inclusions helpful in making a diagnosis of this disorder. The finding of such inclusions in lung biopsy specimens, however, has been distinctly rare. We report herein the ultrastructural findings in the case of a young woman with active systemic LE showing tubuloreticular inclusions within lung and kidney biopsy specimens.     

硫氮酮对失血性休克犬多器官超微结构及脂质过氧化的影响

采用犬失血性休克输血复苏模型，观察了硫氮酮（Ｄｉｌｔｉａｚｅｍ，ＤＺ）对休克犬肝、肾、心、肺４个器官组织超微结构及组织中丙二醛（ＭＤＡ）含量和黄嘌呤氧化酶（ＸＯ）活性的影响。结果显示：ＤＺ可使休克犬各器官组织超微结构的损伤性改变明显减轻；ＤＺ处理组各器官组织中ＭＤＡ含量和ＸＯ活性均显著低于对照组。表明ＤＺ对失血性休克犬多个器官具有保护作用，该效应与其抑制ＸＯ活性和氧自由基介导的脂质过氧化反应有关。     

Glomerular mesangial and endothelial cell swelling following temporary renal ischemia and its role in the no-reflow phenomenon.

The purpose of these experiments was to define changes in glomeruli of rats kidneys which could account for a marked reduction of blood flow immediately following temporary ischemia. After the renal artery had been clamped for 60 minutes, mesangial and endothelial cellular swelling was sufficiently severe to compress and trap intracapillary erythrocytes, obstruct capillary lumens, and prevent reflow of an isotonic carbon suspension. Perfusion of saline solution before the renal arteries were clamped washed blood from the kidney, and the resulting cellular swelling alone was not sufficient to block capillary lumens. Erythrocyte trapping did not occur after 15 minutes of ischemia. Swelling of glomerular and tubular epithelial cells produced some extrinsic capillary compression that could contribute to erythrocyte trapping, but this appeared to be of minor importance in producing the no-reflow phenomenon.     

IMMUNOFLUORESCENCE STAINING IN UNFIXED OR FIXED RENAL BIOPSY SPECIMENS FROM PATIENTS WITH DIABETIC NEPHROPATHY

Immunofluorescence staining in unfixed or fixed renal biopsy specimens were evaluated in nine patients with diabetic nephropathy in order to elucidate if immunofluorescence staining is applicable in fixed renal tissues in such patients. Renal biopsy specimens were embedded in gelatin or paraffin matrix. Renal biopsy specimens embedded in paraffin matrix were digested with 0.05% protease. Immunofluorescent studies of kidney tissues were performed by staining with FITC-labeled heavy chain specific anti-human IgG, IgA, IgM, acute phase reactant (APR) proteins such as α-1-anti-trypsin (αl-AT), haptoglobin (Hpt) and β-lipoprotein (β -Lp) antisera, and then examined with a fluorescent microscope. Linear and nodular deposition of IgG, IgA, IgM, α 1-AT, Hpt, and β -Lp were observed in the glomerular capillary walls of the renal specimens embedded in paraffin matrix. The staining patterns in specimens embedded in paraffin matrix was similar to that embedded in gelatin matrix. There was no significant difference in the intensity or distribution of IgG, IgM, αl-AT, and β-Lp deposition among the two different conditions of immunofluorescence in patients with diabetic nephropathy. It was suggested that immunofluorescence staining in renal biopsy specimens embedded in paraffin matrix after digestion with protease is useful for the evaluation of IgG, IgM, APR proteins, and β -Lp in glomeruli from patiens with diabetic nephropathy.     

Intravascular "mulberry-like" bodies: morphological, immunohistochemical, and ultrastructural observations of an incidental finding caused by autolysis?

Intravascular "mulberry-like" bodies in a stillborn female infant with moderate maceration are reported. The histogenesis of these structures is discussed based on light-microscopic, immunohistochemical and ultrastructural findings. No demonstrable causal relation between the intravascular lesions and fetal death was found, the cause of death being attributed to intrauterine asphyxia. It is concluded, that intravascular "mulberry-bodies" most likely represent artifacts due to red blood cell autolysis.     

Endometrial cell dissemination at diagnostic hysteroscopy: a prospective randomized cross-over comparison of normal saline and carbon dioxide uterine distension.

The incidence of pelvic spreading of endometrial cells at diagnostic hysteroscopy was studied comparing the two distension media carbon dioxide (CO(2)) and normal saline (N/Saline). Thirty patients requiring laparoscopy and hysteroscopy were included in this study, the main indication for surgery being subfertility. Hysteroscopy was performed using both CO(2) and N/Saline distension on each patient, the order of the distension media being randomly allocated. Samples of peritoneal fluid were aspirated from the pouch of Douglas before and after hysteroscopy with each distension medium, and the specimens were investigated cytologically for the presence of endometrial cells. Endometrium was present in 2/30 (6.7%) peritoneal aspirates before and in 15/60 (25%) collected after the hysteroscopies. There was no major difference between liquid or gaseous distension, transtubal reflux of endometrial cells occurring in 7/30 (23.3%) and in 8/30 (26.7%) hysteroscopies respectively. Positive peritoneal cytology was observed significantly more often in patients who were in the proliferative phase of the menstrual cycle [9/14 (64.3%) versus 0/11, P < 0.004]. In conclusion, transtubal dissemination of endometrium occurs in about one quarter of patients, irrespective whether N/Saline or CO(2) is used for uterine distension; there is no advantage to using gaseous distension for hysteroscopy when investigating high-risk cases for endometrial malignancy.     

Modulation of proximal tubular hydraulic conductivity by peritubular capillary oncotic pressure

Fluid absorption from the proximal tubular lumen is probably a multifactorial process. Earlier studies from our laboratory have indicated that a transepithelial hydrostatic and oncotic pressure difference may be the driving force for as much as 30% of the reabsorbed fluid. During saline volume expansion proximal tubular reabsorption declines and the present experiments were undertaken to investigate whether this reduction could be caused by changes in the passively driven flux component. The hydraulic conductivity was therefore determined from the reabsorptive rate in split oil droplets with normal and high hydrostatic pressure gradients across the wall, at the same time as the peritubular capillary net-work was perfused with solutions containing a colloid of high or low concentration. In the reabsorption experiments the split oil droplet radius was measured and in a separate series of experiments the relationship between droplet radius and pressure was determined; this was found to be 7.3 mmHg pressure increase per 1 μm increase in radius. The increase in the rate of reabsorption from the droplets due to increased intraluminal hydrostatic pressure was 1.02±0.13 nl/min/mm tubular length when a solution with a high colloid concentration was perfused through the capillary net-work, compared with 0.41=0.11 nl/min/mm tubular length when a low colloid containing solution was used for perfusion. The hydraulic conductance in the proximal tubular wall at high colloid perfusion was calculated to be 0.54 nl/min mm mmHg while at a low capillary colloid oncotic pressure it was significantly lower 0.025 nl/min mm mmHg. This drop in hydraulic conductance might be one factor responsible for the decline in fluid absorption in animals exposed to saline volume expansion.     

Ultrastructural changes of kidney in Schistosoma mansoni-infected mice

Schistosomiasis is the second threatening parasitic disease after malaria and among Schistosoma spp., Schistosoma mansoni (S. mansoni) affects about 100 million people in tropic regions in Africa and South America. The current study was carried out to investigate ultrastructural changes of the kidney in mice infected with cercariae of S. mansoni, in which 20 Swiss albino mice of 60-day-old were assigned into two groups (10 each). Control group received 1 ml normal saline by intraperitoneal route. Model group were intraperitoneally infected with 1 ml normal saline containing 40 cercariae of S. mansoni/mouse. After 60 days of infection, specimens from the kidneys of both control and infected mice were obtained and processed for transmission electron microscopy (TEM) examination. The main ultrastructural changes were observed in both glomeruli and tubules. Glomerular findings included irregular thickening and splitting of the glomerular basement membrane (GBM), flattening and effacement of the foot processes of podocytes, and proliferation of mesangial cells. Tubular changes were in the form of swelling, atrophy and vacuolation of tubular epithelial cells, and presence of autophagic vacuoles. In conclusion, adopting TEM shows a number of ultrastructural changes in the kidneys of mice infected with cercariae of S. mansoni, most notably thickening and splitting of GBM and flattening and effacement of foot processes of podocytes and tubular autophagic vacuoles. These changes are still unraveled well in the literature.     

Immunofluorescence staining in unfixed or fixed renal biopsy specimens from patients with diabetic nephropathy

Immunofluorescence staining in unfixed or fixed renal biopsy specimens were evaluated in nine patients with diabetic nephropathy in order to elucidate if immunofluorescence staining is applicable in fixed renal tissues in such patients. Renal biopsy specimens were embedded in gelatin or paraffin matrix. Renal biopsy specimens embedded in paraffin matrix were digested with 0.05% protease. Immunofluorescent studies of kidney tissues were performed by staining with FITC-labeled heavy chain specific anti-human IgG, IgA, IgM, acute phase reactant (APR) proteins such as alpha 1-anti-trypsin (alpha 1-AT), haptoglobin (Hpt) and beta-lipoprotein (beta-Lp) antisera, and then examined with a fluorescent microscope. Linear and nodular deposition of IgG, IgA, IgM, alpha 1-AT, Hpt, and beta-Lp were observed in the glomerular capillary walls of the renal specimens embedded in paraffin matrix. The staining patterns in specimens embedded in paraffin matrix was similar to that embedded in gelatin matrix. There was no significant difference in the intensity or distribution of IgG, IgM, alpha 1-AT, and beta-Lp deposition among the two different conditions of immunofluorescence in patients with diabetic nephropathy. It was suggested that immunofluorescence staining in renal biopsy specimens embedded in paraffin matrix after digestion with protease is useful for the evaluation of IgG, IgM, APR proteins, and beta-Lp in glomeruli from patients with diabetic nephropathy.