Incidence and risk factors of osteomyelitis in adult and pediatric systemic lupus erythematosus: a nationwide, population-based cohort study.

OBJECTIVE: The objective of this paper is to investigate the incidence rate, risk factors and outcome of osteomyelitis among patients with systemic lupus erythematosus (SLE). MATERIALS AND METHODS: We conducted a cohort study using data for patients enrolled in the Taiwan National Health Insurance Database from 2000 to 2012. Patients with SLE and age- and sex-matched controls without SLE were enrolled. Primary endpoint was the first occurrence of osteomyelitis. Risks of osteomyelitis in SLE patients were analyzed with Cox proportional hazards regression models, including age, sex, comorbidities and medications. RESULTS: Among 24,705 SLE patients (88.4% women, mean age 35.8 years) with a median follow-up of 9.1 years, 386 patients had osteomyelitis. The incidence rate ratio (IRR) of osteomyelitis in the SLE group vs the control group was 8.52 (95% confidence interval (CI) 7.24-10.05). The SLE group had higher incidence rates of osteomyelitis than the control group, especially in pediatric subgroups (IRR 41.1 95% CI 18.57-107.35). Compared to controls, SLE patients experienced osteomyelitis at a younger age (42.3 vs 58.1 years) but did not have an increased risk of mortality (hazard ratio 0.7; 95% CI 0.21-2.38). Age >60 years, male gender, malignancy within five years, prior bone fracture and higher daily prednisolone dose (>7.5 mg) cumulatively for >180 days increased risk for osteomyelitis. CONCLUSIONS: SLE patients have a higher IRR of osteomyelitis than controls. Pediatric and elder SLE patients, patients with a history of bone fracture, malignancy within five years and higher-dose glucocorticoid use have a higher risk of osteomyelitis and should be carefully monitored.

BCAS2 promotes prostate cancer cells proliferation by enhancing AR mRNA transcription and protein stability.

BACKGROUND: We showed previously that breast carcinoma amplified sequence 2 (BCAS2) functions as a negative regulator of p53. We also found that BCAS2 is a potential AR-associated protein. AR is essential for the growth and survival of prostate carcinoma. Therefore we characterised the correlation between BCAS2 and AR. METHODS: Protein interactions were examined by GST pull-down assay and co-immunoprecipitation. Clinical prostate cancer (PCa) specimens were evaluated by immunohistochemical assay. AR transcriptional activity and LNCaP cell growth were assessed by luciferase assay and MTT assay, respectively. RESULTS: BCAS2 expression was significantly increased in PCa. BCAS2 stabilised AR protein through both hormone-dependent and -independent manners. There are at least two mechanisms for BCAS2-mediated AR protein upregulation: One is p53-dependent. The p53 is suppressed by BCAS2 that results in increasing AR mRNA and protein expression. The other is via p53-independent inhibition of proteasome degradation. As BCAS2 can form a complex with AR and HSP90, it may function with HSP90 to stabilise AR protein from being degraded by proteasome. CONCLUSIONS: In this study, we show that BCAS2 is a novel AR-interacting protein and characterise the correlation between BCAS2 and PCa. Thus we propose that BCAS2 could be a diagnostic marker and therapeutic target for PCa.

Increased serum fibroblast growth factor-23 and decreased bone turnover in patients with systemic lupus erythematosus under treatment with cyclosporine and steroid but not steroid only.

SUMMARY: In patients with systemic lupus erythematosus (SLE), low bone mineral density (BMD) is associated with increased age, prolonged disease, low body mass index (BMI), and overlap with rheumatoid arthritis (RA). Elevated fibroblast growth factor (FGF)-23 in cyclosporine A (CsA) users with SLE are associated with decreased active vitamin D and osteocalcin. INTRODUCTION: The objective of this study was to investigate the steroid and CsA effect on bone metabolism and serum FGF-23 in SLE patients. METHODS: Seventy-two SLE patients and 10 age- and sex-matched healthy individuals underwent blood tests for bone metabolic biomarkers and FGF-23, and lumbar spine dual-energy X-ray absorptiometry for BMD. RESULTS: Comparisons between patients and controls were made in premenopausal women/men younger than 50 years and postmenopausal women/men older than 50 years separately. SLE patients had more frequent low Z-score (≤-2.0, 8.5 vs. 0%), osteopenia (-2.5

Acute respiratory distress syndrome in a pregnant woman with systemic lupus erythematosus: a case report.

When the disease activity of systemic lupus erythematosus (SLE) is controlled appropriately, a pregnant woman who has lupus is able to carry safely to term and deliver a healthy infant. While the physiology of a healthy pregnancy itself influences ventilatory function, acute pulmonary distress may decrease oxygenation and influence both mother and fetus. Though respiratory failure in pregnancy is relatively rare, it remains one of the leading conditions requiring intensive care unit admission in pregnancy and carries a high risk of maternal and fetal morbidity and mortality, not to mention the complexity caused by lupus flare. We report a case of SLE complicated with lupus pneumonitis and followed by acute respiratory distress during pregnancy. Though there is a high risk of maternal and fetal morbidity and mortality, maternal respiratory function improved after cesarean section and treatment of the underlying causes. The newborn had an extremely low birth weight but was well at discharge.

Sustainable Participation in Community Health Programs to Promote a Healthy Lifestyle and Prevent and Protect against Dementia among Rural Taiwanese Middle-Aged and Older Adults.

BACKGROUND: Resource and economic constraints limit access to health care in rural populations, and consequently, rates of chronic illnesses are higher in this population. Further, little is known about how rural populations adopt active and healthy lifestyle behavior for dementia prevention. OBJECTIVES: This study aimed to explore the impact of modification in lifestyle behaviors on changes in cognitive function among middle-aged and older adults living in a rural area of Taiwan. DESIGN: In this prospective longitudinal study, changes in lifestyle and cognitive function were compared between the experimental and control groups. SETTING: Six rural community care stations were randomly cluster sampled in southern Taiwan. PARTICIPANTS: A total of 155 participants were enrolled and classified into two groups according to their community activity participation rate (CAPR). The control group (n=68) had a CAPR < 1x/month, and the experimental group (n=87) had a CAPR ≥ 1x/month. MEASUREMENTS: Cognitive function of the participants was measured using the MMSE scale. Lifestyle behaviors were measured using a self-designed questionnaire based on the Transtheoretical Model. RESULTS: From 2018-2020, the experimental group successfully maintained a healthy lifestyle. The MMSE score in the experimental group was significantly higher in the 3rd year than that in the control group (25.37 vs 22.56, p < 0.001). CONCLUSIONS: Sustainable community participation and adopting a healthy lifestyle could effectively maintain the cognitive function of the study participants. However, more needs to be done to support rural older adults to maintain a healthy diet and control their weight.

Deep lobe parotid abscess with facial nerve palsy: a case report.

Acute suppurative sialadenitis mostly occurs in the parotid gland, while parotid abscesses principally arise in the superficial lobe. However, facial nerve palsy, secondary to parotid abscess, is rare. Predisposing factors for the ductally ascending infection are dehydration, xerogenic drugs and salivary gland diseases associated with ductal obstruction or reduced saliva secretion. Obstruction of Stensen's duct and diminished production of saliva are regarded as the promoting factors. Painful swelling of the preauricular region and cheek is the most familiar symptom of acute suppurative parotitis. The most common pathogens associated with acute bacterial infection are Staphylococcus aureus and anaerobes. We report a rare case of deep lobe parotid abscess with facial nerve palsy. Aside from adequate fluid hydration, good oral hygiene and treatment with empiric parenteral antibiotics, surgical treatment with drainage can provide a remedy for this disease.

Carboxy terminally truncated forms of ribophorin I are degraded in pre-Golgi compartments by a calcium-dependent process.

Two COOH terminally truncated variants of ribophorin I (RI), a type I transmembrane glycoprotein of 583 amino acids that is segregated to the rough portions of the ER and is associated with the protein-translocating apparatus of this organelle, were expressed in permanent HeLa cell transformants. Both variants, one membrane anchored but lacking part of the cytoplasmic domain (RL467) and the other consisting of the luminal 332 NH2-terminal amino acids (RI332), were retained intracellularly but, in contrast to the endogenous long lived, full length ribophorin I (t 1/2 = 25 h), were rapidly degraded (t 1/2 less than 50 min) by a nonlysosomal mechanism. The absence of a measurable lag phase in the degradation of both truncated ribophorins indicates that their turnover begins in the ER itself. The degradation of RI467 was monophasic (t 1/2 = 50 min) but the rate of degradation of RI332 molecules increased about threefold approximately 50 min after their synthesis. Several pieces of evidence suggest that the increase in degradative rate is the consequence of the transport of RI332 molecules that are not degraded during the first phase to a second degradative compartment. Thus, when added immediately after labeling, ionophores that inhibit vesicular flow out of the ER, such as carbonyl cyanide m-chlorophenylhydrazone (CCCP) and monensin, suppressed the second phase of degradation of RI332. On the other hand, when CCCP was added after the second phase of degradation of RI332 was initiated, the degradation was unaffected. Moreover, in cells treated with brefeldin A the degradation of RI332 became monophasic, and took place with a half-life intermediate between those of the two normal phases. These results point to the existence of two subcellular compartments where abnormal ER proteins can be degraded. One is the ER itself and the second is a non-lysosomal pre-Golgi compartment to which ER proteins are transported by vesicular flow. A survey of the effects of a variety of other ionophores and protease inhibitors on the turnover of RI332 revealed that metalloproteases are involved in both phases of the turnover and that the maintenance of a high Ca2+ concentration is necessary for the degradation of the luminally truncated ribophorin.

O-glycosylation of intact and truncated ribophorins in brefeldin A-treated cells: newly synthesized intact ribophorins are only transiently accessible to the relocated glycosyltransferases.

Ribophorins I and II are type I transmembrane glycoproteins of the ER that are segregated to the rough domains of this organelle. Both ribophorins appear to be part of the translocation apparatus for nascent polypeptides that is associated with membrane-bound ribosomes and participate in the formation of a proteinaceous network within the ER membrane that also includes other components of the translocation apparatus. The ribophorins are both highly stable proteins that lack O-linked sugars but each contains one high mannose N-linked oligosaccharide that remains endo H sensitive throughout their lifetimes. We have previously shown (Tsao, Y. S., N. E. Ivessa, M. Adesnik, D. D. Sabatini, and G. Kreibich. 1992. J. Cell Biol. 116:57-67) that a COOH-terminally truncated variant of ribophorin I that contains only the first 332 amino acids of the luminal domain (RI332), when synthesized in permanent transformants of HeLa cells, undergoes a rapid degradation with biphasic kinetics in the ER itself and in a second, as yet unidentified nonlysosomal pre-Golgi compartment. We now show that in cells treated with brefeldin A (BFA) RI332 molecules undergo rapid O-glycosylation in a multistep process that involves the sequential addition of N-acetylgalactosamine, galactose, and terminal sialic acid residues. Addition of O-linked sugars affected all newly synthesized RI332 molecules and was completed soon after synthesis with a half time of about 10 min. In the same cells, intact ribophorins I and II also underwent O-linked glycosylation in the presence of BFA, but these molecules were modified only during a short time period immediately after their synthesis was completed, and the modification affected only a fraction of the newly synthesized polypeptides. More important, these molecules synthesized before the addition of BFA were not modified by O-glycosylation. The same is true for ribophorin I when overexpressed in HeLa cells although it is significantly less stable than the native polypeptide in control cells. We, therefore, conclude that soon after their synthesis, ribophorins lose their susceptibility to the relocated Golgi enzymes that effect the O-glycosylation, most likely as a consequence of a conformational change in the ribophorins that occurs during their maturation, although it cannot be excluded that rapid integration of these molecules into a supramolecular complex in the ER membrane leads to their inaccessibility to these enzymes.

Long-range attractive force between hydrophobic surfaces observed by atomic force microscopy.

There is evidence from atomic force microscopy for a long-range attractive force between hydrophobic surfaces that is virtually identical to that observed with the surface forces apparatus. This force is present in the nonaqueous solvent ethylene glycol. A possible molecular mechanism involves in-plane polarized domains of solid-like monolayers adsorbed on mica, and a theoretical model has been developed that accounts for many of the observations.

Cell growth with trans fatty acids is affected by adenosine 3',5'-monophosphate and membrane fluidity.

Two positional isomers (9 and 11) of trans octadecenoates did not support growth on glucose of an Escherichia coli mutant that requires unsaturated fatty acids. However, the trans fatty acids provided sufficient fluidity to produce much higher cell yields when the concentration of adenosine 3',5'-monophosphate was raised. The effectiveness of the trans acids rose from 0 to 1 cell per femtomole to 15 to 20 cells per femtomole as the concentration of adenosine 3',5'-monophosphate was increased. The corresponding cis positional isomers supported high yields (35 to 40 cells per femtomole) independent of supplementation. The enhanced growth with adenosine 3',5'-monophosphate supplementation is not due to an increased uptake and incorporation of the trans isomers relative to the cis isomers, since the 9-trans isomer was incorporated more rapidly than the 9-cis isomer into the membrane phospholipids under all growth conditions and represented 21 +/- 2 mole percent of the acids. The finding that cells growing with trans fatty acid isomers have a higher requirement for adenosine 3',5'-monophosphate may indicate that some fatty acids can alter the metabolic regulation normally exerted by the cyclic nucleotide.

ErbB4 (JM-b/CYT-1)-induced expression and phosphorylation of c-Jun is abrogated by human papillomavirus type 16 E5 protein.

Human papillomavirus type 16 E5 (HPV-16 E5) is a highly hydrophobic membrane protein with weak-transforming activity, which is associated with ErbB4 receptor in HPV-16-infected cervical lesions. Presently, we investigated the transforming mechanisms of E5 involving ErbB4 signaling. Firstly, we report a role for ErbB4 (JM-b/CYT-1) receptor that activates c-jun gene expression and phosphorylating at Ser63 and Ser73 of the c-Jun protein in ligand-independent and Ras-c-jun NH(2)-terminal kinase-dependent pathway. Secondly, we show that HPV-16 E5 protein can form a complex with ErbB4 via binding to the extracellular and transmembrane domains of ErbB4 (JM-b/CYT-1). When co-expressing HPV-16 E5 and ErbB4 in cells, E5 can abrogate ErbB4-induced c-Jun protein expression and phosphorylation resulted in increasing cell proliferation compared to ErbB4-expressing cells. The interaction between of HPV-16 E5 and ErbB4 provides more insight into the mechanisms of HPV-16 E5 transformation induction.

PEDF induces p53-mediated apoptosis through PPAR gamma signaling in human umbilical vein endothelial cells.

OBJECTIVE: Pigment epithelial-derived factor (PEDF) is a potent anti-angiogenic factor whose effects are partially mediated through the induction of endothelial cell apoptosis. The pathway mediating endothelial cell apoptosis has not been fully established. Here we investigated the participation of peroxisome proliferator-activated receptor gamma (PPARgamma) and p53 in the apoptosis of human umbilical vein endothelial cells (HUVECs). METHODS AND RESULTS: HUVECs pretreated with either PPARgamma antagonist or PPARgamma small interfering RNA (siRNA) suppressed PEDF-induced apoptosis as determined by TUNEL assay, annexin V-FITC/PI staining, and cleavage of procaspase-8, -9, -3. PEDF sequentially induced PPARgamma and p53 expression as observed in immunoblotting and immunofluoresence assays. PEDF also increased the transcriptional activity of PPARgamma as evident from electromobility shift assays, and p53 as determined by the phosphorylation and acetylation of p53 and the induction of Bax. The induction of p53 by PEDF was abolished by either PPARgamma antagonist or PPARgamma siRNA. PEDF-mediated HUVEC apoptosis and cleavage of procaspases were significantly attenuated by p53 siRNA. CONCLUSIONS: Our observations indicate that PEDF induces HUVECs apoptosis through the sequential induction of PPARgamma and p53 overexpression. With the growing interest in anti-angiogenesis as a novel approach to cancer therapy, defining the mechanism of PEDF-mediated HUVEC apoptosis may facilitate the development of new therapeutics.

The involvement of active DNA synthesis in camptothecin-induced G2 arrest: altered regulation of p34cdc2/cyclin B.

Cell cycle arrest in G2 phase is a common response to a variety of DNA-damaging agents. The coupling between DNA damage and G2 arrest was studied in synchronized HeLa cells using camptothecin, a highly specific inhibitor of topoisomerase I that damages DNA through the formation of reversible topoisomerase I-DNA cleavable complexes. Brief camptothecin treatment of early S-phase HeLa cells caused arrest at G2 phase and abolished the activation of p34cdc2 protein kinase. Both tyrosine dephosphorylation of p34cdc2 and cyclin B accumulation were altered. These cell cycle-dependent changes were not observed when DNA replication was inhibited by aphidicolin during the brief camptothecin treatment. Our results suggest that to produce G2 arrest, active DNA synthesis is required at the time of camptothecin treatment, as was previously shown for camptothecin-induced cytotoxicity. Furthermore, our results suggest that the interaction of the replication fork with DNA damage may ultimately trigger altered regulation of p34cdc2/cyclin B, leading to cell cycle arrest at the G2 phase.

Interaction between replication forks and topoisomerase I-DNA cleavable complexes: studies in a cell-free SV40 DNA replication system.

The extreme S-phase-specific cytotoxicity of camptothecin has been shown to involve active DNA replication. To investigate the role of DNA replication in camptothecin cytotoxicity, we have studied the interaction between the DNA replication machinery and the topoisomerase I-camptothecin-DNA ternary cleavable complex in a cell-free SV40 DNA replication system. The formation of topoisomerase I-camptothecin-DNA-cleavable complexes on the replication template efficiently and irreversibly inhibited DNA replication. Two aberrant forms of replication products were produced whose abundance varied with the concentrations of exogenously added topoisomerase I and camptothecin. At low concentrations of topoisomerase I and camptothecin, the major aberrant DNA replication product was close-to-unit-length-linear DNA, while at higher concentrations the predominant product was close-to-dimer-size-linear DNA. Analysis of these aberrant replication products has suggested a "collision" model in which the interaction between an advancing replication fork and a topoisomerase I-camptothecin-DNA-cleavable complex results in irreversible arrest of the replication fork and the formation of a double-strand DNA break at the fork. Concomitant with fork arrest and fork breakage, the reversible cleavable complex was converted into a topoisomerase I-linked DNA break. We propose that one or several of these events triggers S-phase-specific cell killing and G2-phase cell cycle arrest.

Audit of phlebotomy turnaround time in a private hospital setting.

An audit on the turnaround time (TAT) of a hospital phlebotomy service was undertaken to assess whether or not the existing service standards can satisfy the needs and expectations of both the external and internal customers. A job request survey form was designed and implemented in June 2005 to be used by the day shift to record the number of phlebotomy cases and the corresponding TAT. The success rate and complaints related to phlebotomy were also recorded. Phlebotomists provided the data for this study on an honor system basis. Out of 2,118 test requests received by the laboratory, 1,867 (88.1 percent) were phlebotomy requests. Approximately 62 phlebotomy requests were recorded, on average, per day shift. The average time, expressed in mean +/- standard deviation (SD) needed for response, arrival, and job completion was 7.4 +/- 1.5 minutes, 5.6 +/- 1.6 minutes, and 10.4 +/- 2.4 minutes respectively, with an average overall TAT of 23.4 +/- 4.1 minutes per phlebotomy request. The success rate at first phlebotomy attempt was 97 percent, and only one complaint was received during the audit period. This study may help hospital management identify possible bottlenecks that delay phlebotomy TAT, thus improving service standards in this area.

Thermodynamic and phase characterization of phosphatidylethanolamine and ganglioside GD1a mixtures.

By employing diphenylhexatriene steady-state fluorescence anisotropy, pyrenedecanoic acid excimer formation, and high sensitivity scanning calorimetry we have demonstrated that the liposomes containing phosphatidylethanolamine (PE) and various mole fractions of ganglioside GD1a had a gel-liquid crystalline phase transition between 15 and 25 degrees C. Calorimetric measurements indicated that these phase transitions were broad and centered between 17 and 21 degrees C. The enthalpy change of the transition was linearly dependent on the ganglioside concentration up to 10.0 mol% and plateaued between 11.4-16.2 mol%. The high enthalpy change (37 kcal/mol of GD1a added into the PE bilayer) indicates the existence of PE-GD1a complex structure in the liposomal membrane. It is proposed that semi-fluid domains containing six PE and one ganglioside molecule are present in the PE-GD1a membranes at temperatures above gel-liquid crystalline phase transition. The Sendai virus induced leakage of PE-GD1a liposomes has been investigated by using an entrapped, self-quenching fluorescent dye, calcein. The leakage rate was dependent on the mole fraction of ganglioside GD1a and was maximal at 6.3 mol%. Arrhenius plots of the leakage rates showed breaks in the 20-25 degrees C temperature range, which correspond to the gel-liquid crystalline phase transition of the target liposomes. These data suggest that the rate of Sendai virus-induced leakage can be regulated via fluidity modulation by changing the PE to GD1a ratio at constant temperatures.

Monitoring Chinese hamster ovary cell culture by the analysis of glucose and lactate metabolism.

Monitoring cell growth is crucial to the success of an animal cell culture process that can be accomplished by a variety of direct or indirect methodologies. Glucose is a major carbon and energy source for cultured mammalian cells in most cases, but glycolytic metabolism often results in the accumulation of lactate. Glucose and lactate levels are therefore routinely measured to determine metabolic activities of a culture. Typically, neither glucose consumption rate nor lactate accumulation rate has a direct correlation with cell density due to the changes in culture environment and cell physiology. We discovered that although the metabolic rate of glucose or lactate varies depending on the stages of a culture, the cumulative consumption of glucose and lactate combined (Q(GL)) exhibits a linear relationship relative to the integral of viable cells (IVC), with the slope indicating the specific consumption rate of glucose and lactate combined (q(GL)). Additional studies also showed that the q(GL) remains relatively constant under different culture conditions. The insensitivity of the q(GL) to process variations allows a potentially easy and accurate determination of viable cell density by the measurement of glucose and lactate. In addition, the more predictable nature of a linear relationship will aid the design of better forward control strategies to improve cell culture processes.

Cloning and expression of the genes associated with lipid metabolism in Tsaiya ducks.

Sterol regulatory element binding protein 1 (SREBP1) drives the expression of several lipogenic genes, whereas SREBP2 dictates the expression of every gene involved in cholesterolgenesis in mammals. In the current study, we cloned the cDNA fragments for SREBP1, SREBP2, fatty acid synthase (FAS), 3-hydroxy-3-methylglutaryl-CoA reductase (HMG-CoA reductase), and very low density apolipoprotein-II (apoVLDL-II), the genes associated with lipid metabolism. Fifteen ducks immediately before the first egg was laid (18 wk old) and 15 ducks from the same population at an egg production rate of 80% were killed. Total RNA was extracted from liver and used to amplify the targeted genes by reverse transcription-PCR and screening of a cDNA library. The sequence data showed that Tsaiya duck SREBP1, SREBP2, FAS, and HMG-CoA reductase were highly homologous to that of chicken. Tsaiya duck SREBP1 mRNA was expressed in adipose tissue, cardiac muscle, skeletal muscle, liver, and ovary. The SREBP2 mRNA concentration was highest in liver and ovary. Concentrations of FAS and HMG-CoA reductase mRNA were high in liver and lower in other tissues. The apoVLDL-II mRNA was specifically expressed in the liver. The differences between mRNA concentrations of SREBP1, SREBP2, and FAS in the livers of laying and prelay ducks were not significant. However, the concentrations of hepatic HMG-CoA reductase and apoVLDL-II mRNA were higher in the laying ducks than in prelay ducks. Therefore, laying may affect particular aspects of lipid metabolism, especially biochemical pathways that involved apoVLDL-II and HMG-CoA reductase.

A Golgi-related structure remains after the brefeldin A-induced formation of an ER-Golgi hybrid compartment.

Brefeldin A (BFA) has previously been shown to block protein transport from the endoplasmic reticulum (ER), to cause the redistribution of Golgi components to the ER, and to change profoundly the morphology of the Golgi apparatus. In order to quantitate the effects of this drug on the morphology of the ER and the Golgi apparatus in HeLa cells, the numerical, surface and volume densities of these organelles were determined by stereological means. We found that in cells treated with BFA (5 micrograms/ml) clusters of vesicles and tubules, often located near transitional elements of the ER, replaced the Golgi apparatus. The numerical density of these clusters in cells treated with BFA for 30 min or 4.5 h is similar to that of Golgi complexes and Golgi-related clusters in control cells. The surface density of the vesicles and tubules contained in these clusters is about 50% of that represented by Golgi elements in control cells. Concomitantly, a corresponding increase in the surface density of the ER-Golgi hybrid compartment was observed. This hybrid compartment contained Golgi-specific enzymes effecting modifications of N-linked oligosaccharides and the transfer of O-linked sugars. Antibodies recognizing different subcompartments of the Golgi apparatus or the intermediate compartment, labeled vesicles and tubules of the Golgi-related clusters. Applying low doses of BFA allowed for the dissection of the disassembly of the Golgi apparatus into at least two phases. At very low doses (10-20 ng/ml) the numerical density of vesicles in the clusters increased up to 4-fold above control, while the surface density did not markedly change, suggesting that vesiculation of the Golgi cisternae had occurred. Fusion of Golgi elements with the ER seemed to occur only at doses of BFA higher than 20 ng/ml. Contrary to observations on other cell types, removal of BFA from HeLa cell cultures resulted in a rather slow reformation (1-2 h) of the Golgi complex, which allowed us to observe several intermediate stages in this process. During this time period an ER was restored which no longer contained Golgi-specific O-glycosylation functions. Our results demonstrate that BFA does not simply cause the disappearance of the Golgi apparatus by fusion with the ER, but instead clusters of vesicles and tubules remain that contain Golgi-specific markers.