Lymphocyte characterization of decidua basalis spiral arteries with acute atherosis in preeclamptic and normotensive pregnancies.

Uteroplacental acute atherosis (AA) is a common spiral arterial lesion in preeclampsia, characterized by intramural foam cells, fibrinoid necrosis, and a perivascular immune cell infiltrate. A clear definition of this infiltrate is lacking. Therefore, our aim was to characterize lymphocytes in pre-defined zones regarding spiral arteries with or without AA, from preeclamptic and normotensive pregnancies. Lymphocytes were characterized in decidua basalis samples (n = 91), previously evaluated for AA, around spiral arteries in three pre-defined zones; 1) intramural, 2) perivascular and 3) interstitial. Adjacent serial sections were immunostained to identify different T-cell populations (CD3+, CD8+, FOXP3+), and NK-cells (CD56+). CD3+CD8- T-cells were also identified. These were presumed to be largely CD4+ T-cells. AA was associated with significantly higher intramural CD3+ cell concentrations in Zone 1, in both normotensives and preeclamptics. In preeclamptics only, this difference extended into Zone 2. Similar results were observed for CD3+CD8- cells. AA was also associated with increased intramural CD8+ concentration; however, the number of cells was low. Regulatory T-cells (FOXP3+) were generally scarce or absent in all pre-defined zones. Although intramural NK-cells (CD56+) were scarce, the intramural concentration was significantly lower in spiral arteries with AA compared to without AA in preeclamptics. Our main finding was that CD3+CD8-FoxP3- T-cells were associated with AA. We therefore suggest that T-cells, of a non-regulatory CD4+ subtype, could be involved in the formation of spiral artery AA in the decidua basalis. Whether AA gives rise to, or is partly mediated by increased T-cell concentration around the lesions, remains to be determined.

Effects of explant size on epithelial outgrowth, thickness, stratification, ultrastructure and phenotype of cultured limbal epithelial cells.

PURPOSE: Transplantation of limbal stem cells is a promising therapy for limbal stem cell deficiency. Limbal cells can be harvested from either a healthy part of the patient's eye or the eye of a donor. Small explants are less likely to inflict injury to the donor site. We investigated the effects of limbal explant size on multiple characteristics known to be important for transplant function. METHODS: Human limbal epithelial cells were expanded from large versus small explants (3 versus 1 mm of the corneal circumference) for 3 weeks and characterized by light microscopy, immunohistochemistry, and transmission electron microscopy. Epithelial thickness, stratification, outgrowth, ultrastructure and phenotype were assessed. RESULTS: Epithelial thickness and stratification were similar between the groups. Outgrowth size correlated positively with explant size (r = 0.37; P = 0.01), whereas fold growth correlated negatively with explant size (r = -0.55; P < 0.0001). Percentage of cells expressing the limbal epithelial cell marker K19 was higher in cells derived from large explants (99.1±1.2%) compared to cells derived from small explants (93.2±13.6%, P = 0.024). The percentage of cells expressing ABCG2, integrin ß1, p63, and p63α that are markers suggestive of an immature phenotype; Keratin 3, Connexin 43, and E-Cadherin that are markers of differentiation; and Ki67 and PCNA that indicate cell proliferation were equal in both groups. Desmosome and hemidesmosome densities were equal between the groups. CONCLUSION: For donor- and culture conditions used in the present study, large explants are preferable to small in terms of outgrowth area. As regards limbal epithelial cell thickness, stratification, mechanical strength, and the attainment of a predominantly immature phenotype, both large and small explants are sufficient.

Effect of Transportation on Cultured Limbal Epithelial Sheets for Worldwide Treatment of Limbal Stem Cell Deficiency.

Limbal stem cell deficiency can be treated with transplantation of cultured human limbal epithelial cells (LEC). It can be advantageous to produce LEC in centralized labs and thereafter ship them to eye clinics. The present study used transport simulations of LEC to determine if vigorous shaking during transport altered the viability, morphology and phenotype during a 4 day-long storage of LEC with a previously described serum-free storage method. Inserts with LEC cultured on amniotic membranes were sutured to caps inside air-tight containers with generous amounts of 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES)-buffered minimal essential medium (MEM). The containers were distributed among the following testing conditions: 6 hours with full containers, 36 hours with full containers, 36 hours with container three quarters full of medium, and 36 hours with container full of medium containing a shear-protecting agent (Pluronic-F68). Compared to stored, but non-transported controls, no statistically significant changes in viability and immunohistochemical staining were observed. The epithelial sheets remained intact. However, an air-liquid interface in the containers reduced the number of desmosomes and hemi-desmosomes compared to the controls. In conclusion, cultured LEC sheets appear to endure vigorous shaking for at least 36 hours if the container is full.

Febrile Illness and Intravenous Antimalarial Treatment at a District Hospital in Cameroon.

BACKGROUND: Malaria is one of the most common causes of mortality worldwide. The World Health Organization (WHO) has recently recommended that treatment should be guided by a laboratory diagnosis. The aim of this study is to explore patient and health care factors associated with intravenous antimalarial treatment of malaria test-negative patients at a district hospital in a malaria endemic area. STUDY DESIGN: A cross-sectional study of 91 patients admitted for intravenous antimalarial treatment was done at a district hospital in northern Cameroon in July and August 2010. Socio-cultural and clinical factors were studied in relation to quality blood smear results. RESULTS: Thirty-two per cent of all intravenous antimalarials were administered to patients with a negative malaria test. Test negative patients older than 40 years of age received significantly more often intravenous antimalarials than the youngest patients (OR = 7.9, 95% CI = 1.9-32.4, p = 0.004). Few differential diagnoses were identified in the study population, and patients older than 30 years of age had malaria less often than the youngest patients (OR = 0.3, 95% CI = 0.08-0.9 and OR = 0.09, 95% CI = 0.02-0.4). CONCLUSION: This study supports previous reports of over-diagnosis and treatment of malaria in endemic areas. The results suggest that differential diagnoses are important, especially in adults with febrile illnesses to ensure the correct diagnosis and treatment. Further studies are needed to explore the findings and to develop strategies to improve the management of malaria and its differential diagnoses.

The acceptability of volunteer, repeat blood donations in a hospital setting in the Adamaoua region of Cameroon.

BACKGROUND AND OBJECTIVES: The knowledge of factors that may influence blood donation in Cameroon is limited. The objectives of this study are to assess the characteristics of previous and potential blood donors by exploring the religious beliefs, and knowledge and understanding of blood donations among individuals present at a district hospital. MATERIALS AND METHODS: Forty-nine in-depth, semi-structured interviews were conducted among consenting, randomly selected 18 years or older community members present at a district hospital in the Adamaoua region during October and November 2011. RESULTS: Ninety-eight per cent (48/49) of the individuals present at this district hospital had heard of blood transfusions. Forty-seven per cent (23/49) had not previously been asked to donate blood; however, 94% (44/47) said that they would donate if given the opportunity. Thirty-three per cent (16/49) had previously donated blood to family members or for replacement, and 81% of these said they would repeat donations. The majority of both donors and non-donors were motivated to donate blood for altruistic reasons. CONCLUSION: The findings suggest that community members present at this district hospital in Cameroon may be recruited for repeat blood donations. Although the altruistic motivation to donate blood suggests that donors could be recruited from a district hospital population, targeted information about blood donations and accessible blood transfusion services need to be put in place. The study may add to the understanding of the preconditions for blood donations and the possibility to establish sustainable blood transfusion services in the Adamaoua region in Cameroon.

A study of blood transfusion services at a district hospital in Malawi.

BACKGROUND AND OBJECTIVES: Severe anaemia is an important cause of mortality in developing countries. However, few studies have explored the use of and possibilities for blood transfusion services. The aims of this study are to explore the use of blood transfusion services at a hospital in sub-Saharan Africa and to assess the quality of the transfusion services according to WHO guidelines. MATERIALS AND METHODS: Patient age, gender, haemoglobin (Hb) level, diagnosis, hospital department and replacement donations were recorded for all blood transfusions administered at a district hospital in Malawi in January 2010. The laboratory equipment and procedures were scored according to WHO guidelines. RESULTS: The mean Hb of transfused patients was 4·8 g/dl. Fifty-seven per cent (59/104) of the transfusions were given to children diagnosed with malaria, and 17% (18/104) were given to pregnant women. During the study period, blood was in stock and available for transfusion within 1 h of requisition. The equipment and procedures at this hospital met the main criteria for an adequate WHO stage of development. CONCLUSION: In contrast to the advanced transfusion medicine in developed nations, our findings highlight the persistent and urgent need for life-saving blood transfusions in especially young children and pregnant women in Africa. The results indicate that blood transfusion services adapted to local conditions may be a realistic solution for providing safe blood products in developing countries. Serious challenges, such as HIV transmission and sustainable organization of low-risk blood donations should be addressed to assure access to safe blood products.

A new, clinically oriented, unifying and simple placental classification system.

OBJECTIVE: At present there is no internationally accepted, clinically easy understandable, comprehensive morphological placental classification. This hampers international benchmarking and comparisons, and clinical research. STUDY DESIGN: Internationally published criteria on morphological placental pathology were collected, standardized and focused into a comprehensive diagnosis category system. The idea was to create a clinically relevant placental pathology scheme related to major pathological processes. A system of nine main diagnostic categories (normal placenta included) was constructed. Pathologists and obstetricians discussed the mutual understanding of the wording in the reporting. The previously published diagnostic criteria were merged, structured and standardized. Through an interobserver correlation study on 315 placentas from intrauterine deaths and 31 controls (placentas from live births) the microscopic criteria in this classification system were tested on user-friendliness and reproducibility. RESULTS: The clinical feedback has been very positive, focusing on the understandability and usefulness in patient follow-up. The interobserver agreement in the microscopic correlation study was in general good. The differences in agreement mainly reflected the degree of preciseness of the microscopic criteria, exemplified by excellent correlation in diagnosing acute chorioamnionitis. Maternal and fetal circulatory disorders need grading criteria and studies are needed to get more insight and clinical correlations of villitis and maturation disorders. CONCLUSION: The clinically oriented, unifying and simple placental pathology classification system may work as a platform for standardization and international benchmarking. Further research is needed to define diagnostic criteria in staging and grading of some main diagnostic categories.

Effects of serum-free storage on morphology, phenotype, and viability of ex vivo cultured human conjunctival epithelium.

The use of amniotic membrane (AM) represents one of the major developments in ocular surface reconstruction. However, in a study on patients with primary pterygium, transplantation of AM with ex vivo expanded human conjunctival epithelial cells (HCjE) promoted earlier epithelialization than AM alone. We previously showed that cultured human limbal epithelial cells maintain their morphology, phenotype, and viability for one week when stored at 23°C. The current study investigates the feasibility of storing HCjE in HEPES-MEM and Optisol-GS at 23°C for 4 and 7 days, respectively. The five experimental groups were analyzed by light microscopy, immunohistochemistry, transmission electron microscopy, and a viability assay. The ultrastructural integrity of cultured HCjE was well preserved following 4 days of storage, however, 7 days of storage resulted in some loss of cell-cell contacts and epithelial detachment from the amniotic membrane. The number of microvilli in cultured HCjE not subjected to storage was 2.03±0.38 microvilli/µm. In comparison, after 4 and 7 days of HEPES-MEM storage this number was 1.69±0.54 microvilli/µm; P=0.98 and 0.89±1.0 microvilli/µm; P=0.28, respectively. After Optisol-GS storage for 4 and 7 days, the mean number of microvilli was 1.07±1.0 microvilli/µm; P=0.47 and 0.07±0.07 microvilli/µm; P=0.03, respectively. The number of cell layers in cultured HCjE not subjected to storage was 4.4±0.3 cell layers, as opposed to 4.0±0.9 cell layers; P=0.89 after 4 days of HEPES-MEM storage and 2.8±0.6 cell layers; P=0.01 after 7 days of storage in HEPES-MEM. The number of cell layers after 4 and 7 days of storage in Optisol-GS was 3.7±0.2 cell layers; P=0.46 and 3.4±0.4 cell layers; P=0.18, respectively. The expression of markers for undifferentiated cells (ΔNp63α, ABCG2 and p63), proliferating cells (Ki67 and PCNA), goblet cells (Ck7 and MUC5AC), stratified squamous epithelial cells (Ck4), and apoptotic cells (caspase-3) in cultured HCjE appeared to be unchanged after 4 and 7 days of HEPES-MEM and Optisol-GS storage. The percentage of viable cells in cultured HCjE not subjected to storage (91.4%±3.2%) was sustained after 4 and 7 days of storage in HEPES-MEM (94.1%±4.5%; P=0.99 and 85.1%±13.7%; P=0.87, respectively) as well as after 4 and 7 days of storage in Optisol-GS (87.7%±15.2%; P=0.97 and 79.8%±15.7%; P=0.48, respectively). We conclude that cultured HCjE may be stored for at least 4 days in serum-free conditions at 23°C while maintaining the phenotype and viability. HEPES-MEM appears to be comparable to Optisol-GS for serum-free storage with preservation of the ultrastructure for at least 4 days.

Consideraciones sobre un caso singular de mutilación digestiva / Considerations an a singular case of digestive mutilation

El contenido de este trabajo se ajusta a la presentación de un caso notable que expone dos particularidades primordiales: 1) paciente sometida sucesivamente a: colectomía total, colecistectomía, gastrectomía total y ulteriormente a resección de los dos tercios superiores del recto; y 2) que haya sido factible practicar una evaluación clínica y paraclínica a más de tres décadas de la primera intervención quirúrgica, posibilitando de este modo estimar y debatir distintos ángulos de enfoque del caso en cuestión.

The remarcable case presented on this paper has two special characteristics: 1) the patient received, sequentially, a total colectomy, a cholecystectomy, a total gastrectomy and a resection of the two upper thirds of the rectum; and 2) the fact that it was feasible to make a clinical and paraclinical assesment more than three decades after the first surgery, making it possible to consider and to debate the different approaches towards this particular case.

Homeobox gene distal-less 3 is expressed in proliferating and differentiating cells of the human placenta.

DLX3, a member of the large homeobox gene family of transcription factors, is necessary for normal placentation. Targeted deletion of dlx3 in mouse resulted in embryonic death due to placental failure. This study demonstrates the presence of DLX3 mRNA expression in human first trimester and term placental tissue, cultured trophoblast-like cell lines and in isolated primary villous and extravillous trophoblast cells. Using an ovine polyclonal antibody, the spatial distribution was identified for DLX3 in human placental tissues, trophoblast cell lines and in freshly isolated primary trophoblast cells. A 50 kDa immunoreactive DLX3 protein was detected in the human placenta, in trophoblast cell lines and in primary trophoblast cells. Nuclear expression for DLX3 was observed in villous cytotrophoblasts, syncytiotrophoblast and extravillous cytotrophoblast in the proximal regions of the cytotrophoblast cell columns in first trimester placental tissues. Immunoreactivity was also detected in few stromal cells and microvascular endothelial cells surrounding the fetal capillaries. In the first trimester placental bed, DLX3 expression was predominantly observed in the cytoplasm of the endovascular and interstitial trophoblasts. We conclude that the cellular expression of DLX3 was extensive in the human placenta and propose that DLX3 may play an important role in normal placental development.