Metabolic and immunological consequences of vitamin D deficiency in obese children.

Numerous studies highlighted the link between vitamin D deficiency and cardiovascular, autoimmune, metabolic diseases, and obesity. However, a clear role of vitamin D in these disorders is still unknown. Vitamin D deficiency in children can be a potential risk factor for developing diseases at a later age. Early prevention and vitamin D supplementation should become a public health priority. This review highlights the clinical implications of vitamin D deficiency in adults and children with obesity.

Pathogenic role of antibodies against monomeric C-reactive protein in tubulointerstitial nephritis and uveitis syndrome.

Antibodies against monomeric C-reactive protein, which is a target antigen expressed both in kidney tubules and uveal cells, have been recently detected in patients with active tubulointerstitial nephritis and uveitis syndrome. We report the case of an 65-year-old woman with acute renal failure caused by biopsy-proven tubulointerstitial nephritis and the onset of uveitis 21 months later. The expression of monomeric C-reactive protein in kidney oligobiopsy was confirmed by immunohistochemical staining using mouse monoclonal antibody against human monomeric C-reactive protein. The levels of antibodies against monomeric C-reactive protein were 117% of the reference during the flare and 22% during the remission of the disease. The difference in the levels of antibodies against monomeric C-reactive protein during flare and remission, and above all positive biopsy staining, supports their pathogenic role in this disease.

Serological studies to determine the occurrence of Johne's disease and mycoplasma infection in the Northern-East Polish population of European bison (Bison bonasus).

A serological study of twenty three European bison (Bison bonasus) derived from Northern-East Poland for the seroprevalence of Mycobacterium avium subsp. paratuberculosis, Mycoplasma bovis, Mycoplasma mycoides subsp. mycoides SC, Mycoplasma agalactiae and Mycoplasma capricolum subsp. capripneumoniae was conducted. Only specific antibodies to M. bovis were detected in two animals (8.7%) which were connected with the clinical signs and macroscopic anatomopathological lesions.

The abnormal superficial radial artery does not restrict the successful creation of hemodialysis forearm arteriovenous fistula.

Anatomical variations of the radial artery are of clinical importance in end-stage renal disease patients awaiting creation of native arteriovenous fistula for hemodialysis. As radial-cephalic direct wrist fistula is a vascular access of choice, atypical localization of the distal part of the radial artery may lead to the false assumption of severe atherosclerotic lesions and prevent creation of such an access, despite good vessel conditions and convenient surgical approach. We present 7 patients with radial artery variations. In 5 patients with superficial radial artery, radial-cephalic direct wrist access was created. One patient, due to an anomaly misdiagnosis, had radial-cephalic fistula created on the contra lateral wrist. In the patient with hypoplastic radial artery brachial-basilic upper arm transposition was created.

Beliefs about back pain and pain management behaviours, and their associations in the general population: A systematic review.

Previous mass media campaigns have aimed to influence how people manage back pain, with mixed success. Campaigns should target beliefs which are related to the behaviours they aim to change. This systematic review brings together research that has measured the prevalence of beliefs about back pain in the general population and factors associated with these beliefs, including future pain-related outcomes. Five databases were searched up until April 2017. Quantitative studies which reported a measure of agreement with a belief about back pain, cross-sectional associations, or associations between beliefs and future outcomes were eligible. Eligibility was assessed and data extracted independently by two authors. Results were tabulated and narratively synthesized. Nineteen studies from 10 countries were eligible (median study n [IQR] = 990.5 [524.75-2387.5]). Beliefs were measured using eight questionnaires and 57 stand-alone items. Beliefs about back pain's negative consequences were common across countries and populations, whereas most samples did not hold fear-avoidance beliefs. Beliefs about back pain's consequences were associated with pain and disability, but only one study investigated this specific relationship prospectively. No studies investigated whether beliefs are associated with future pain management behaviours. Agreement with certain beliefs (e.g. about negative consequences) was associated with sociodemographic characteristics (e.g. older age) and poorer self-rated health. Interventions may benefit from targeting beliefs about the perceived negative consequences of back pain in these populations. However, future research should explore how beliefs prospectively influence the management of back pain. SIGNIFICANCE: This review brings together studies which have assessed the prevalence of beliefs about back pain, and factors associated with holding them. It highlights that whether or not these beliefs represent important determinants of how people manage pain remains unknown.

Endoscopic Islet Autotransplantation Into Gastric Submucosa-1000-Day Follow-up of Patients.

BACKGROUND: Total pancreatectomy and autologous transplantation of pancreatic islets is a treatment option for patients with severe pain due to chronic pancreatitis. In the standard procedure, pancreatic islets are isolated and subsequently administered into the portal vein. In the case of patients with a history of thrombosis or at risk of thrombosis, this route of administration is not viable. Animal studies conducted in our department led to the development of a technique of endoscopic islets transplantation into the gastric submucosa. In 2013 and 2014, the first human autologous transplant procedures were performed. The objective of this study was to present the results of a 3-year follow-up of these patients. METHODS: Two pancreatectomies were performed in our department, the first in 2013 and another in 2014, along with subsequent autologous transplantation of pancreatic islets into the gastric submucosa. RESULTS: Both patients had been diagnosed previously with diabetes, and both had endogenous islet activity detected. Peptide C concentration after pancreatectomy and before pancreatic cell transplantation was 0.1 ng/mL. After the transplantation, peptide C concentrations for the 2 patients were 0.8 and 0.5 ng/mL on day 7, 1.2 and 0.6 ng/mL on day 30, 1.3 and 0.8 ng/mL on day 180, 1.1 and 0.7 ng/mL on day 360, and 3.0 and 0.6 ng/mL at 3 years, respectively, after transplantation. The pain symptoms resolved in both cases. CONCLUSION: Pancreatic islets may survive in the gastric wall. Endoscopic submucosal transplantation may present an alternative for the management of patients who cannot undergo a classic transplantation procedure.

B Cell Activating Factor (BAFF) in Long-term Kidney Transplant Recipients Is Not a Prognostic Marker for Allograft Dysfunction or Survival.

OBJECTIVE: B cell activating factor (BAFF) has been shown to play a role in B cell survival, maturation, and activation, and has been linked with renal transplant outcome. BAFF signaling has been associated with plasmablast survival, anti-HLA immunization, and loss of graft function. We aimed to analyze the interplay between BAFF, memory B cells, and plasmablasts in relation to allograft function in long-term kidney transplant (KTx) recipients and their anti-HLA sensitization. MATERIALS AND METHODS: This study included 70 long-term KTx recipients on standard immunosuppression 15 ± 6 years post transplantation (44 stable, 26 chronic allograft dysfunction, CAD) and 25 healthy volunteers. CD19+ B cells, memory B cells (CD19+CD27+), and plasmablasts (CD19+CD24-CD27++CD38++) were enumerated with flow cytometry. BAFF serum level and anti-HLA antibodies were assessed by Luminex bead arrays. RESULTS: We found no difference in BAFF levels between KTx recipients and controls (median, interquartile range: 1.67, 1.40-1.97 vs 1.78, 1.63-1.93 ng/mL, P = .478) and no correlation between BAFF level and cell counts. Recipients presented lower plasmablast count than controls (22.5, 8-57 vs 79, 48-166 cells/mL, P < .001). There was a positive correlation between estimated glomerular filtration rate and plasmablasts (rs = 0.30, P = .013) in recipients. Cell populations and BAFF were not related to the presence of anti-HLA antibodies. None of the parameters investigated was related to deterioration of allograft function during the 2-year follow-up. CONCLUSION: BAFF serum level is not related to anti-HLA sensitization, circulating memory B cells, plasmablast count, or allograft function. Circulating plasmablasts are associated with current allograft function but are not prognostic for future course.

Expression of PBMC TLR4 in Renal Graft Recipients Who Experienced Delayed Graft Function Reflects Dynamic Balance Between Blood and Tissue Compartments and Helps Select a Problematic Patient.

Both Toll-like receptor 4 (TLR4) and monocytes focus stimuli, causing them to contribute differently to chronic injury of a transplanted kidney. AIM: The aim of our study was to determine if TLR4 monocyte is a diagnostic tool and possibly a target for therapeutic intervention. MATERIALS: We studied 143 kidney transplant (KT) patients (88 male, 55 female; 50.3 ± 12.8 years); median was 10.4 post KT, follow-up was 11.4 months, and 46 patients had delayed graft function (DGF+) history. Control group (38 healthy volunteers) had monocyte mRNA-TLR4 expression (TLR4ex). DGF+ were divided by median of TLR4ex (-0.1034) into 2 groups: low-TLR4 expression (L-TLR4ex) and high-TLR4 expression (H-TLR4ex). RESULTS: We showed that in comparison with DGF-, the DGF+ had much lower TLR4ex, and worse KT function both currently (TLR-day) (serum creatinine [sCr] P = .002; estimated glomerular filtration rate [eGFR] P = .001) and post follow-up (sCr P = .006; eGFR P = .005). The DGF+ with L/H-TLR4ex comparison showed no differences in TLR-day KT function but did show differences in post follow-up (sCr P = .01; eGFR P = .02; ΔeGFR% P = .001). Regression analysis showed an association between recipient age, tacrolimus concentration, and uremic milieu (ie, TLR-day sCr and GFR with TLR4ex). Reverse regression analysis indicated an association of TLR4ex (especially L/H-TLR4ex) with post follow-up parameters of KT function and numeric/qualitative measures of change. CONCLUSION: DGF affects the fate of a graft. Within a several months after transplantation, TLR4ex of peripheral blood mononuclear cells declines in DGF patients. Low LR4ex in patients with DGF+ is associated with poor prognosis for the efficiency of the KT. In patients with DGF+, the proper selection of immunosuppression (tacrolimus dosing) is very important. Higher concentrations of tacrolimus may improve prognosis. The analysis of TLR4ex change may be a useful parameter for the real assessment of immunosuppression efficacy. It is important for transplanted organ function that peripheral blood mononuclear cells effectively leave circulation and remain in the graft.

Pretransplant Immune Interplay Between Donor and Recipient Influences Posttransplant Kidney Allograft Function.

BACKGROUND: Renal transplant candidates present immune dysregulation caused by chronic uremia, and deceased kidney donors present immune activation induced by brain death. Pretransplant donor and recipient immune-related gene expression were examined in the search for novel predictive biomarkers crosslinking recipient and donor pretransplant immune status with transplant outcome. MATERIALS AND METHODS: This study included 33 low-risk consecutive renal transplant recipients and matched deceased donors. The expression of 29 genes linked to tissue injury, T-cell activation, cell migration, and apoptosis were assessed in postreperfusion kidney biopsies, as well as 14 genes in pretransplant peripheral blood of the kidney recipients. Gene expression was analyzed with real-time polymerase chain reaction on custom-designed low-density arrays. RESULTS: Donor MMP9 expression was related to delayed graft function occurrence (P = .036) and short term kidney allograft function (14th day rs = -0.44, P = .012; 1st month rs = -0.46, P = .013). Donor TGFB1 expression was associated with short- and long-term graft function (14th day rs = -0.47, P = .007; 3rd month rs = -0.63, P = .001; 6th month rs = -0.52, P = .010; 12th month rs = -0.45, P = .028; 24th month rs = -0.64, P = .003). Donor TGFB1 expression was not related to donor age (rs = 0.32, P = .081), which was also an independent factor influencing the outcome. Recipient gene expression was not related to graft function but determined the acute rejection risk. Recipient IFNG and, to a lesser extent, IL18 expression were protective against acute rejection (area under the curve [AUC] 0.84, P < .001, and AUC 0.79, P < .001, respectively). CONCLUSION: Kidney transplant outcome depends on the interplay between donor-related immune factors, which mostly affect allograft function and recipient immune milieu, influencing an alloreactive response.

Endothelin A Receptors Expressed in Renal Blood Vessels of Renal Transplant Patients Are Connected With Acute Tubular Necrosis or Antibody-Mediated Rejection.

BACKGROUND: The role of non-HLA antibodies named antiendothelin A receptor antibodies is potentially significant but not established. The significance of the endothelin A receptor (ETAR) and its expression in renal biopsy has not been defined. We decided to evaluate the presence and relevance of ETARs in renal transplant biopsy for cause. The aim of our study was to evaluate the immunoreactivity of the ETAR and its significance in patients who had a renal transplant biopsy due to deterioration of transplant function (biopsy for cause) with detailed characterization of staining in small and intermediate arteries of renal transplant biopsies. METHODS: Immunohistochemical expression of ETARs was analyzed in 162 renal transplant biopsies. Microscopic evaluation of ETAR expression (polyclonal antibody) was performed on paraffin sections. ETAR expression was analyzed in renal blood vessels (small and intermediate arteries) based on three-step scale. RESULTS: We analyzed 154 patients who had renal allograft biopsy between 6 days and 24 years (median 597 days) after transplantation. Positive staining of ETAR in small and intermediate arteries was noticed in 9 patients. Among these patients, 4 had early biopsies (<3 months after transplantation), all developed acute tubular necrosis, and 1 developed additionally acute humoral rejection. Further, 4 patients had late biopsy (1-8 years after transplantation) and all developed characteristics of antibody mediated rejection. Lastly, 1 patient had no characteristic changes in the biopsy 4 months after transplantation. Graft loss 1 year after biopsy was higher in patients who were ETAR-positive but statistical significance was not achieved. CONCLUSIONS: The expression of endothelin receptors in renal blood vessels (small and intermediate arteries) seems to be important in diagnosis of damage during acute tubular necrosis and antibody-mediated rejection.

Histopathological Relevance of Angiotensin II Type 1 Receptor in Renal Transplant Biopsy.

The occurrence of anti-angiotensin II type 1 receptor (AT1R) antibodies is thought to be a risk factor for transplant injury, but the relationship of AT1R to graft loss in renal transplantation has not been assessed. The aim of our study was to evaluate the expression of AT1R and its relationship with graft loss in patients who had a renal transplant biopsy for cause. METHODS: AT1R immunoreactivity was analyzed in 170 renal transplant biopsies. Immunohistochemical evaluation of AT1R expression was performed on 4 µm-thick paraffin sections mounted on silanized slides. AT1R expression was analyzed in 5 compartments: 1. glomeruli, 2. renal blood vessels (small and intermediate arteries), 3. peritubular capillaries, 4. tubular epithelium, and 5. interstitium based on a 3-step scale. RESULTS: Initially we checked 170 consecutive samples of biopsies for the immunoreactivity of the AT1R. The study finally included 118 renal transplant patients in 1-year observation after the biopsy. The renal allograft biopsy was performed between 6 days and 24 years after transplantation and the diagnosis was based on Banff criteria. We observed positive immunostaining of AT1R in tubular epithelium in 26.3% (42/118) of patients. A total of 7 patients had staining assessed as 2 and 35 as 1. One year post-biopsy graft loss in the AT1R (+) patients was 35.7 % (15/42) compared to 14.5% (11/76) in the AT1R (-) group (P = .008). CONCLUSIONS: The expression of AT1R in tubular epithelium of the biopsy for cause was associated with significantly higher graft loss. The relevance of AT1R should be considered for better transplant immunological risk assessment.

The Impact of Functioning Arteriovenous Fistula on Blood Pressure Control and Renal Allograft Function.

INTRODUCTION: The prevalence of hypertension in renal graft recipients is high. It was postulated that central arteriovenous anastomosis may significantly reduce blood pressure. This preliminary study evaluates the impact of functioning arteriovenous fistula (AVF) on blood pressure control and renal allograft function. MATERIALS AND METHODS: One hundred sixty-two previously hemodialyzed kidney transplant recipients (108 males, 54 females, aged 52.7 ± 13.2 years, mean 6.9 ± 5.1 years after transplantation), who had scheduled visits in the first two weeks of March 2015, were included in the study. The recipients were divided into two groups depending on AVF function (65 AVF+ and 97 AVF-). RESULTS: Functioning AVF was more prevalent in males than females (47.2 % vs 25.9 %, P = .009). Both groups presented similar allograft function despite the fact that interval from transplantation to examination day in the AVF+ group was significantly shorter than in the AVF- group (5.2 ± 5.3 vs 8.1± 4.5 years; P < .001). The mean systolic blood pressure (135.0 ± 17.0 vs 138.7 ± 14.1 mm Hg, P = .13) was similar in both study groups, but diastolic blood pressure in the AVF+ group was lower than in the AVF- group (80.0 ± 7.0 vs and 83.7 ± 9.2 mm Hg, P = .006). The proportion of patients with diastolic blood pressure >80 mm Hg was significantly higher in patients without functioning AVF (35 % in the AVF- group vs 20 % in the AVF+ group, P= .038). In multivariate analysis, AVF presence was the only factor significantly influencing a diastolic blood pressure with odds ratio 0.43 (95% CI 0.19-0.99, P = .048), which supports AVF as a potentially positive influence on blood pressure control. CONCLUSIONS: The presence of AVF in renal transplant recipients was associated with a slight decrease in diastolic blood pressure without clear effect on renal function.

Islets Allotransplantation Into Gastric Submucosa in a Patient with Portal Hypertension: 4-year Follow-up.

BACKGROUND: Islets transplantation is an established treatment method for patients suffering from brittle diabetes with hypoglycemia unawareness. The standard implantation technique is through the portal vein into the liver. In case of liver diseases or portal hypertension, finding an extra-hepatic site is recommended. There have been attempts to perform islets transplantations into muscles and into the gastric submucosa. OBJECTIVE: The aim of this study is to show a 4-year follow-up of allotransplantation into gastric submucosa in a case of portal hypertension observed during the procedure of islets infusion. PATIENTS AND METHODS: A 36-year-old woman with complicated diabetes for over 30 years was selected to receive simultaneous islets and kidney transplantation. The patient underwent an unsuccessful simultaneous pancreas and kidney transplantation 2 years earlier in another transplantation center. The patient's daily insulin requirement was 60 IU, which corresponded to 1.15 IU/kg of body weight. The HbA1c level was 7.4%. C-peptide levels, both fasting and stimulated, were 0.01 ng/mL. On December 7, 2013, the patient received transplanted kidney and islets procured from the same donor. Only 124,000 islets equivalents (IEQ) were isolated (2400 IEQ/kg body weight). Islets were suspended in 300 mL of Ringer's solution along with albumin, antibiotics, and heparin. After infusing 100 mL of the islets suspension into the portal vein, pressure in portal vein increased from 5 mm Hg to 23 mm Hg. Despite stopping the infusion, pressure did not drop after 30 minutes. The decision was made to transplant the reminder of the islets (200 mL) into the gastric wall. RESULTS: No complications were observed after the procedure. Serum creatinine level was 1.6 mg/dL on day 10 and 1.5 mg/dL 4 years after the transplantation. Fasting C-peptide levels were 1.7, 0.65, 0.55, 0.69, 0.68, and 0.2 ng/mL at 1, 3, 6, 12, 18, and 36 months after the transplantation, respectively. HbA1c levels were 5.2, 6.4, 4.7, 5.2, and 5.9% at 3, 6, 12, 18, and 36 months, respectively. The patient's insulin requirement dropped to 15 U/day immediately after transplantation and equaled 20 and 27 U/day at 18 and 48 months after the simultaneous islet and kidney transplantation, respectively. CONCLUSION: Allotransplantation of islets into the gastric wall may be a safe alternative in cases of contraindications for transplantation into the portal vein.

Risk factors for glucose metabolism disorders after kidney transplantation with uneventful course.

A wide range of glucose metabolic disorders (GMDs) often arise after renal transplantation that predispose to graft dysfunction, infections, and cardiovascular disease. This study evaluated the risk factors for GMDs among 50 patients including 30 males and overall mean age 44.9 +/- 12.1 years. All 50 subjects displayed normal glucose tolerance tests pretransplantation and no family history of diabetes. They were selected from the 99 consecutive patients transplanted from April 2005 to January 2006 based upon uneventful posttransplantation course, without rejection episodes or hepatitis C virus (HCV) infections. The study concentrated on risk factors originating during the dialysis period. Even in this selected group, the risk of posttransplant GMD development was high (28%). Patients with GMDs showed significantly worse renal function at 1 month after transplantation (serum creatinine concentration: 1.70 +/- 1.67 mg/dL in the GMD group vs. 1.44 +/- 0.96 mg/dL in the group without GMDs [P = .027] and eGFR, 56.68 +/- 22.70 mL/min/1.73 m(2) versus 71.29 +/- 27.37 mL/min/1.73 m(2), respectively, [(P = .099)]. In a logistic regression model, a statistically significant difference between the groups was shown only for cold ischemia time (P = .037). In the logistic regression model with two independent variables, statistical significance was observed (P = .038) for body mass index at the time of transplantation. In this model, a lower pretransplant serum insulin concentration showed an influence that bordered on significance (P = .074). This study confirmed that the etiology of GMD after kidney transplantation is multifactorial, and at least in part connected with the pre-transplantation period.

Weight reduction in renal transplant recipients program: the first successes.

INTRODUCTION: Overweight and obesity in kidney graft recipients, both at transplantation and further on, are connected with the development of complications of metabolic syndrome. Hypertension, diabetes, and atherosclerosis are risk factors for chronic allograft nephropathy, shortened graft function, and lower recipient life expectancy. The aim of this study was to present the initial results from a weight reduction in renal transplant recipients program. MATERIAL: Thirty-four overweight and obese kidney transplant recipients were enrolled in the study: 9 overweight (26%), 19 obese (55.8%), and 6 morbidly obese (17.6%). The control group encompassed 418 kidney transplant recipients, in whom fluctuations in body mass and body mass index (BMI) were monitored for 56 months. METHODS: During the first visit, we performed an account of dietary habits and anthropometric measurements. At the second visit following a 6-month interval, patients received dietary guidelines based on an analysis of diet questionnaires. RESULTS: Six months after enrollment, despite not having received dietary guidelines during the first visit, only 27% of study subjects and 80% of controls experienced weight gain. CONCLUSIONS: Patients enrolled in the first step of the weight reduction program had no weight nor BMI increase after 6 months. Recipients having experienced body mass increase constituted only 27% of the study group, whereas increase in body mass occurred in 80% of controls. Reducing body mass accretion in kidney transplant recipients should be the target of preventive measures and nonpharmacological therapeutic interventions conducted by qualified personnel. Greater interest by medical personnel in the issue of body mass increase in recipients may be a strong motivating factor for them to undertake weight loss measures.

Posttransplant overweight and obesity: myth or reality?

UNLABELLED: Posttransplant body mass index (BMI) increase in kidney transplant recipients is an underestimated issue, predisposing to morbidity linked with development of polymetabolic syndrome. AIM: The aim of the study was to assess the incidence of overweight and obesity among endstage renal disease patients before and after kidney transplantation. MATERIAL: Four hundred eighteen kidney graft recipients were enrolled in the study which lasted a mean of 56 months. Inhabitants of Lower Silesia (n = 3855) were used as controls. Overweight was defined as BMI between 25 and 30 kg/m(2) and obesity as >30 kg/m(2). METHODS: Mean BMI calculated in 418 patients, both pretransplant and after a 4.5-year observation period was compared with results of the Lower Silesian population. RESULTS: Mean pretransplant BMI in men (n = 242) and women (n = 189) was lower than in controls: men pretransplant BMI 24.3 kg/m(2) versus 25.7 kg/m(2) in the normal population; women, pretransplant BMI 23.17 kg/m(2) versus 25.2 kg/m(2) in the control group respectively. Mean total pretransplant BMI values increased from 23.82 to 25.9 kg/m(2) at last checkup ("last BMI"). A lesser posttransplant BMI increase was noted in men (7%) compared with women (9.6%). Before transplant, overweight or obesity occurred in 38% (n = 157), after a 4.5-year observation period, 65% (n = 232). CONCLUSIONS: Our observations documented that obesity is a widespread issue in kidney graft recipients, affecting two thirds of the population. It should be the target of preventive measures and nonpharmacologic therapeutic interventions.

Immunohistochemical analysis of in vivo patterns of Bak expression, a proapoptotic member of the Bcl-2 protein family.

The in vivo patterns of bak gene expression were determined in human tissues using an immunohistochemical approach. Polyclonal antisera were raised in rabbits against a synthetic peptide corresponding to amino acids 14-36 of the human Bak protein, and were shown to be specific by immunoblot analysis of various human tissues and cell lines. Bak immunoreactivity was detected in a wide variety of cell types and was typically present within the cytosol in a punctuate pattern suggestive of association with intracellular organelles. Consistent with a proapoptotic role for the Bak protein, gradients of Bak protein production were observed in the complex epithelia of the nasopharynx, esophagus, colon, and bladder, with Bak immunointensity being highest in the upper layers and relatively low in the basal portions of these epithelia. Similarly, in the myeloid series of hematopoietic cells, Bak immunoreactivity was strongest in the terminally differentiated granulocytes, with only weak immunostaining occurring in most progenitor cells in the bone marrow. Among the other cell types and tissues with prominent Bak immunostaining were: (a) cardiomyocytes; (b) vascular and visceral smooth muscle cells; (c) basal cells of the prostate glands; (d) myoepithelial cells of the mammary glands; (e) distal convoluted tubules of the kidney; (f) epidermal keratinocytes; (g) enterocytes of the small intestine; (h) Sertoli and Leidig cells of the testes; (i) theca interna cells in the ovary; and (j) adrenal cortex (but not adrenal medulla). Nearly all neurons and glial cells of the central nervous system did not contain immunodetectable Bak protein, whereas sympathetic neurons as well as neurons in dorsal root ganglia and their axons were Bak immunopositive. Most circulating peripheral blood lymphocytes were negative for Bak immunostaining, whereas strong Bak immunoreactivity was found frequently in lymphocytes in the nodes and spleen. Overall, these patterns of bak expression are unique compared to other members of the bcl-2 gene family, and suggest that bak regulates cell death at specific stages of cell differentiation through tissue-specific control of its expression.

Elevated expression of Bcl-X and reduced Bak in primary colorectal adenocarcinomas.

Expression of several members of the BCL-2 family of genes was investigated by immunohistochemical methods in 30 primary colorectal adenocarcinomas and 24 adenomatous polyps. When compared to the intensity observed in adjacent normal mucosal epithelial cells, the intensity of Bcl-X immunostaining was elevated in 18 of 30 (60%) carcinomas (P = 0.0001) and 12 of 24 (50%) adenomatous polyps (P = 0.0001). Immunoblot analysis of five pairs of tumors and adjacent normal colonic tissue indicated marked elevations in the relative levels of the anti-apoptotic Bcl-XL, protein in all cases. In contrast to the increased Bcl-X expression, the intensity of Bcl-2 immunostaining was greater than that of normal colonic mucosa in only 3 of 30 (10%) carcinomas and, in fact, was lower than that of adjacent normal epithelia] cells in 25 (83%) cases (P = 0.0001). Furthermore, the percentage of Bcl-2 immunopositive cells was generally lower in carcinomas than in adenomas (mean +/- SE, 44 +/- 6% versus 73 +/- 5%, respectively; P = 0.001) and in moderately or poorly differentiated tumors than in well-differentiated tumors (39 +/- 6% versus 70 +/- 11%, respectively; P = 0.045). In addition, the proportion of tumors in which the Bcl-2 immunointensity was more than or equal to that of normal colonic mucosa was significantly lower in carcinomas than adenomas (5 of 30 versus 15 of 24, respectively; P < 0.001), suggesting that decreases in Bcl-2 expression represent a later event associated with the progression of colorectal cancers. When compared to that of normal adjacent colonic epithelium, the intensity of Mcl-1 immunostaining was reduced in 20 of 30 (67%) of carcinomas (P = 0.0001) compared to only 1 of 24 adenomas, suggesting that decreases in Mcl-1 expression represent a later event associated with progression from a benign to a malignant phenotype or with transition to a less-differentiated state, because most of the carcinomas evaluated here (25 of 30; 83%) were not well differentiated. The intensity of immunostaining for the pro-apoptotic protein Bak was reduced compared to that of normal mucosal epithelial cells in 27 of 30 (90%) carcinomas and 22 of 24 (92%) adenomas, suggesting that reductions in Bak expression occur early in colorectal tumor progression (P = 0.0001). In contrast, the intensity of immunostaining for the pro-apoptotic protein Bax was not significantly altered in carcinomas; compared to that of normal colonic mucosa, Bax immunointensity was reduced in only 7 of 30 (23%) carcinomas and 3 of 24 (13%) adenomas, and the percentage of Bax immunopositive cells was also not significantly different in any of the histological subgroups. Taken together, these results suggest that expression of Bcl-XL is increased in undifferentiated primary colorectal cancers, often with accompanying reciprocal decreases in the anti-apoptotic proteins Bcl-2 and Mcl-1 and the pro-apoptotic protein Bak, whereas Bax expression is relatively constant. Thus, a shift from expression of the anti-apoptotic proteins Bcl-2 and Mcl-1 to the Bcl-XL protein may occur during progression of colorectal tumors.