[A predictive model based on risk factors for early mortality in patients with newly diagnosed multiple myeloma].

Objective: To investigate risk factors for early mortality (EM) in patients with newly diagnosed multiple myeloma (NDMM) and to build an EM-predictive model. Methods: In a cohort of 275 patients with NDMM, risk factors for EM at 6, 12, and 24 months after diagnosis (EM6, EM12, and EM24, respectively) were determined to establish a model to predict EM. Results: The rates of EM6, EM12, and EM24 were 5.5% , 12.7% , and 30.2% , respectively. The most common cause for EM was disease progression/relapse, accounting for 60.0% , 77.1% , and 84.3% of EM6, EM12, and EM24, respectively. EM6 was associated with corrected serum calcium >2.75 mmol/L and platelet count <100×10(9)/L, whereas risk factors for EM12 included age >75 years, ISS Ⅲ, R-ISS Ⅲ, corrected serum calcium >2.75 mmol/L, serum creatinine >177 µmol/L, platelet count <100×10(9)/L, and bone marrow plasma cell ratio ≥ 60% . In addition to the risk factors for EM12, EM24 was also associated with male sex and 1q21 gain. By multivariate analysis, age >75 years, platelet count <100×10(9)/L, and 1q21 gain were independent risk factors for EM24 but there were no independent risk factors significantly associated with EM6 and EM12. Using a scoring system including these three risk factors, a Cox model for EM24 was generated to distinguish patients with low (score<3) and high (score ≥ 3) risk. The sensitivity and specificity of the model were 20.7% and 99.2% , respectively. Further, an internal validation performed in a cohort of 183 patients with NDMM revealed that the probability of EM24 in high-risk patients was 26 times higher than that in low-risk patients. Moreover, this model was also able to predict overall survival. The median overall survival of patients with scores of 0, 1, 2, 3, 4, and 5 were 59, 41, 22, 17.5, and 16 months, respectively. Conclusion: In the study cohort, the EM6, EM12, and EM24 rates were 5.5% , 12.7% , and 30.2% , respectively, and disease progression or relapse were main causes of EM. An EM24-predictive model built on three independent risk factors for EM24 (age>75 years, platelet count<100×10(9)/L, and 1q21 gain) might predict EM risk and overall survival.

[Epidemiological analysis of cytogenetic abnormalities in patients with newly-diagnosed multiple myeloma: a multi-center retrospective study].

Objective: To analyze the frequency and composition of risk-related cytogenetic abnormalities (CAs) in patients with newly-diagnosed multiple myeloma (NDMM) . Methods: The frequency and composition of risk-related CAs from a cohort of 1 015 Chinese patients with NDMM were determined by interphase fluorescence in situ hybridization (iFISH) , individually or in combination. Results: Of the cohort of 1 015 Chinese patients with NDMM, the frequencies of IgH arrangement, del (13q) /13q14, 1q gain and del (17p) were 54.0%, 46.4%, 46.1% (35.8% and 12. 7% for 3 or more than 3 copies) and 9.9%, respectively. Among 454 patients who had the baseline information for all risk-related CAs [except t (14;20) , which was not covered by the FISH panels performed routinely at all five centers], the frequencies of t (4;14) , t (11;14) or t (14;20) were 14.1%, 11.2% and 4.8%, respectively; of them, 44.3% patients carried 2 or more CAs (28.0%, 13.4% and 2.9% for 2, 3 or ≥4 CAs) ; 83.3%, 95.0% or 68.6% patients with 1q gain, del (17p) or IgH rearrangement had 1 or more additional CA (s) , with del (13q) /13q14 as the most frequently accompanied CA; 57.7% patients carried at least 1 HRCA; the incidences of double-hit (DH) MM (DHMM) (=2 HRCAs) and triple-hit (TH) (THMM) (≥3 HRCAs) were 14.3% and 2.9%, respectively. Conclusions: Our results provided an up-to-date profile of CAs in Chinese NDMM patients, which revealed that approximately 58% patients might carry at least 1 HRCA, and 17% could experience so-called DHMM or THMM who presumably had the worst outcome.

[Adverse effects of double-hit combining ISS-â ¢ stage and 1q gain or del (17p) on prognosis of patients with newly diagnosed multiple myeloma].

Objective: To evaluate the prognostic significance of combining ISS-Ⅲ and high risk cytogenetic abnormalities [HRCAs, including 1q gain/amplification and del (17p) ] in patients with newly-diagnosed multiple myeloma (NDMM) . Methods: The clinical characteristics and relevant variables were retrospectively analyzed in a total of 270 NDMM patients diagnosed between November 2009 and May 2018. ISS-Ⅲ stage and HRCAs [detected by FISH, including 1q gain/amplification and del (17p) ] were defined as risk factors (hit) . Based to the number of hit per case, these patients were divided into four groups carrying 0 to 3 risk factors, respectively. Progress-free survival (PFS) and overall survival (OS) were then analyzed using the Kaplan-Meier estimator. Results: Patients who carried single hit (n=120, 44.4%) had shorter median PFS (23.0 vs 28.9 months; P>0.05) and OS (42.3 vs 53.7 months; P>0.05) than those with no risk factors (n=66, 24.4%) . Of note, the outcome of patients who had two or more risk factors (double/triple, n=84, 31.1%) was much worse than those with either no or one risk factor, indicated by significantly reduced median PFS (14.5 months; HR=1.584, 95%CI 1.082-2.319; P=0.003 for double/triple vs single hit) and OS (18.4 months, HR=2.299, 95%CI 1.485-3.560; P<0.001 for double/triple vs single hit) . Strikingly, patients who had three risk factor (triple hit, n=5, 1.9%) displayed the poorest survival with extraordinarily shorter PFS (0.9-15.1 months) and OS (0.9-18.9 months) compared to those carrying two risk factors (double hit) . Analogous results were obtained when different combinations of ISS stages and HRCAs were analyzed. Conclusion: These results suggest a potential but rather important role of combining multiple (e.g. double or triple) adverse factors determined via the routine ISS staging and FISH detection of cytogenetic abnormalities in risk stratification and prognostic prediction, which might be helpful to identify high risk patients more precisely at diagnosis. It also raised a possibility that a small group of ISS-Ⅲ patients carrying both 1q gain/amplification and del (17p) might represent an "extremely-high risk" subset of MM.

[Prognostic analysis of 182 newly diagnosed multiple myeloma patients with high risk cytogenetic abnormalities].

Objectives: To evaluate the clinical characteristics and prognosis of high risk cytogenetic abnormalities (HRCA) and various combinations of cytogenetic abnormality in patients with newly-diagnosed multiple myeloma (NDMM) . Methods: This retrospective study collected 182 NDMM patients in the First Affiliated Hospital of Jilin University between Nov. 2009 and May 2018. HRCA included 1q+, del (17p) , t (4;14) , and t (14;16) detected by FISH, and non-HRCA included del (13q) , t (11;14) detected by FISH. The clinical characteristics among three groups, including cases who carrying a single HRCA, 1 HRCA in combination with non-HRCA and cases carrying two or more HRCAs (double/triple-hit) were observed. Kaplan-Meier curve was used to analyze both progression-free survival (PFS) and overall survival (OS) for the three groups. Results: The survivals of patients with 1 HRCA in combination with non-HRCA were similar to those with two or more HRCAs (double/triple-hit) , the median PFS (mPFS) was 19.1 m vs 12.1 m (P=0.248) and median OS (mOS) was 29.6 m vs 29.3 m (P=0.774) . Furthermore, the prognosis of these two groups were both inferior to patients with a single HRCA, respectively. (mPFS: 32.2 m, P=0.040, P=0.001; mOS: 42.3 m, P=0.021, P=0.041) . Strikingly, both the mPFS and the mOS of patients with 1 HRCA in combination with non-HRCA (regardless of high risk or not) were significantly shorter than that of cases with a single HRCA (mPFS: 15.1 m vs 32.2 m, HR=2.126, 95%CI 1.176-3.843, P=0.005; mOS: 29.3 m vs 42.3 m, HR=1.442, 95%CI 0.705-2.950, P=0.011) . Conclusion: It is of prognostic significance value for detecting double/triple-hit based on FISH cytogenetics in NDMM.

[The prognostic significance of dynamic monitoring of minimal residual disease (MRD) status in patients with newly-diagnosed multiple myeloma].

Objective: To evaluate the prognostic value of kinetic changes in minimal residual disease (MRD) status, as well as its relationship with risk stratification, therapeutic response and treatment in patients with newly-diagnosed multiple myeloma (MM) . Methods: A total of 135 patients with newly-diagnosed MM were screened, and 105 patients who achieved VGPR or more as the best responses were included into this study. The MRD status was determined by multiparameter flow cytometry (MFC) at multiple intervals after two cycles of treatment until clinical relapse, death, or last follow-up. The statistical methods included Kaplan-Meier analysis, Cox regression, etc. Results: ①In all 135 patients, 57.8% (78/135) patients achieved MRD negativity (MRD(-)) after treatment. In 105 patients who achieved VGPR and thus included in this study, the MRD(-) rate was 72.4% (76/105) , with a median interval of 3 months from starting treatment to achievement of MRD(-) status. ②The 2-year PFS rate of patients with MRD(-) status was significantly higher than that of MRD(+) status (62.2% vs 41.3%, P=0.001) , while MRD persistence (MRD(+)) was an independent factor for poor prognosis (multivariate analysis for PFS: P=0.044, HR=3.039, 95%CI 1.029-8.974) . ③Loss of MRD(-) status (i.e., MRD reappearance) showed inferior outcomes compared with MRD sustained negative ones, the PFS was 18 months versus not reach (P<0.001) and the OS was not reach for both (P=0.002) . ④The 2-year PFS and OS rates of patients with duration of MRD(-)status≥12 months were significantly higher than those of the control group (PFS: 77.7% vs 36.7%, P<0.001; OS: 96.4% vs 57.9%, P<0.001 respectively) . Duration of MRD(-) status was associated with a marked reduction in risk of relapse or death (univariate analysis for PFS: P<0.001, HR=0.865, 95%CI 0.815-0.918; for OS: P=0.001, HR=0.850, 95%CI 0.741-0.915 respectively) . ⑤Moreover, even in patients carrying high-risk cytogenetic abnormalities (CA) or ineligible for ASCT, MRD negativity remained its prognostic value to predict PFS (high-risk CA medianPFS: not reach vs 19 months, P=0.006; ineligible for ASCT medianPFS: not reach vs 25 months, P=0.052 respectively) . ⑥Last, treatment with the bortezomib-based regimens contributed to prolonged MRD(-) duration (median MRD(-) duratio: 25 months vs 10 months, P=0.034) . Conclusion: Our findings supported MRD(+) status as an independent poor prognostic factor in MM patients, which implicated that duration of MRD(-) status also played a significant role in evaluation of prognosis, while loss of MRD(-)status might serve as an early biomarker for relapse. Therefore, monitoring of MRD kinetics might more precisely predict prognosis, as well as guide treatment decision, especially for when to start retreatment in relapsed patients.

[Effect of 1q21 amplification on bortezomib therapeutic response and prognosis of newly diagnosed multiple myeloma patients].

Objective: To investigate the effect of 1q21 amplification (1q) on the therapeutic response and prognosis of bortezomib(Btz) in the treatment of newly diagnosed multiple myeloma (MM) patients. Methods: A total of 180 newly diagnosed MM were included for analyses of clinical characteristics, cytogenetics, objective response rate (ORR), progression-free survival (PFS) and overall survival (OS), retrospectively. Gene expression profiling (GEP) was analyzed using publicly available R2 platform. Results: ① In 180 patients, 1q was found in 51.1% cases. Of them, 174 patients had complete follow-up data, including 88 cases with 1q and 86 without 1q (non-1q). ②Incidence of 1q was positively associated with percentage of IGH rearrangement (72.2%, P=0.017) and 1p deletion (1p) (27.8%, P=0.040). ③ The median PFS was 15.0 and 20.3 months for the 1q group and non-1q group, and the median OS was 29.4 and 44.0 months, respectively. Both PFS and OS of 1q group was significantly shorter than those of the non-1q group (P=0.029 and 0.038, respectively). Multivariate analysis further revealed that 1q was an independent prognostic factor for both PFS (HR=1.910, 95% CI 1.105-3.303, P=0.020) and OS (HR=2.353, 95% CI 1.090-5.078, P=0.029). ④ In 91 evaluable cases with 1q, very good partial remission (VGPR) rate was higher after treatment with Btz than those without Btz (62.1% vs 40.0%, P=0.032). Of note, the patients with 1q who received auto-HSCT after induction with Btz had significantly longer PFS than those without auto-HSCT (19 months vs 13 months, P=0.048). ⑤GEP analysis revealed that 1q21 amplification predominantly up-regulated expression of >50% genes within 1q21 region, and also altered expression of 28% genes in chromosome 1 and 10% genes in whole genome, particularly related to DNA repair and cell cycle. Conclusions: 1q is an independent adverse prognostic factor in patients with newly diagnosed MM. It is often associated with 1p deletion and IGH rearrangement. Patients with 1q respond well to Btz-based regimen, but they fail to gain long-term benefit from this treatment itself. However, auto-HSCT following Btz induction might improve survival of patients with 1q, suggesting a potential strategy to treat this high-risk subset of MM. GEP analysis warrants further attention in understanding the mechanisms underlying the high-risk of 1q.

[A retrospective study of the BiRd regimen in the treatment of relapsed/ refractory multiple myeloma].

Objective: To evaluate efficacy of the BiRd regimen, a combination of clarithromycin, lenalidomide, and dexamethasone, in the treatment of patients with relapsed/refractory multiple myeloma (RRMM) . Methods: Patients with RRMM treated with BiRd between September 11, 2013 and August 1, 2016 at six centers were included to evaluate overall survival rate (ORR) , clinical benefit rate (CBR) , progression-free survival (PFS) , overall survival (OS) , as well as adverse events. Results: Of 30 patients with RRMM, 27 patients were evaluable, and ORR and CBR were 51.9% (14/27) and 66.7% (18/27) respectively, including 1 sCR (3.7%) , 3 CR (11.1%) , 3 VGPR (11.1%) , and 7 PR (25.6%) . In 13 patients with prior Rd, ORR and CBR were 38.5% (5/13) and 61.5% (8/13) respectively, of which 5 patients with ≥MR carried high-risk cytogenetic[ (e.g.17p- or t (4;14) ] together with at least one of other adverse-prognostic cytogenetic (e.g.13q- and/or 1q21+) . In 24 patients with prior bortezomib-based therapy, ORR and CBR were 45.8 and 62.5%, respectively. With a median follow-up time of 14.9 (range 1.0-33.8) months, the median PFS and OS were 12.0 (95%CI 11.6-12.4) and 27.6 (95%CI 15.1-40.1) months, respectively. The BiRd regimen was well tolerated. Conclusion: The BiRd regimen is an effective and safety protocol for RRMM, including those carrying high-risk cytogenetic markers.

[A prospective multi-center trial of non-interventional and observational study of lenalidomide in Chinese patients with multiple myeloma].

Objective: To evaluate the efficacy and safety of lenalidomide in a real-world clinical practice in Chinese patients with multiple myeloma (MM). Methods: It was a prospective, multi-center, observational study. A total of 165 consecutive patients with MM treated with lenalidomide-based regimens were enrolled in 12 hospitals from June 2013 to November 2015. Relevant information was recorded, such as baseline clinical data, cytogenetic abnormalities, treatment regimens, and duration of treatment, safety, and survival. Results: (1)There were 126 relapsed and refractory MM (RRMM) patients, 25 newly diagnosed patients and 19 maintenance patients. The evaluable RRMM patients accounted for 120 cases, among which 74 cases(61.7%) reached the partial response (PR) or above, and a very good partial response (VGPR) in 16 patients (13.3%), a complete response (CR) in 14 cases (11.7%), a strictly complete response (sCR) in 4 cases (3.3%). Thus, a VGPR or above in 34 patients accounted for 28.3%. (2)The median follow-up was 13 months, the median time to progression 12 months. The median survival after receiving lenalidomide was 19 months, and the median overall survival (OS) was 62 months. (3) The univariate analysis in 120 RRMM patients suggested that prognostic factors for significant improvement in PFS included normal karyotype, international staging system (ISS) Ⅰ-Ⅱ, t(4; 14) negative (detected by fluorescence in situ hybridization), non-bortezomib resistance and response to previous regimens. As to OS, non-bortezomib resistance, response to previous regimens and non-primary refractoriness were positive factors. Multivariate analysis showed that the response to previous regimens (PR or better) was an independent good prognostic factor for progress-free survival(PFS), non-bortezomib resistance and non-primary refractoriness for OS. (4) Grade 3 or 4 adverse events that occurred in more than 10% of all enrolled patients were neutropenia (12.7%), leukocytosis(11.5%) and thrombocytopenia (12.7%). Owing to intolerance of toxic side effects, 7 cases withdrew lenalidomide. Conclusions: No matter what combination, regimens containing lenalidomide are effective to RRMM patients with overall response rate 61.7%, a time to progression 12 months and an overall survival 62 months.The toxicity is quite tolerable and manageable. In addition, the response to previous treatment (reached PR or above) is the independent good prognostic factor for PFS, non-bortezomib resistance and non-primary refractoriness for OS. Clinical trail registration: Clinicaltrials.gov, NCT01947309.

Predictors of recent HIV testing in homosexual men in Australia.

OBJECTIVES: To describe time trends and other predictors of recent HIV testing among homosexual men enrolled in behavioural surveillance studies in Australia. METHODS: Repeated cross-sectional studies during the period 1996-2001 in Australian capital cities. Men were recruited from a variety of community-based settings, including gay community outdoor events, sex on premises venues, and social venues. They underwent a brief self-administered questionnaire in which they reported their HIV status, HIV-testing history, sexual behaviour and demographic information. RESULTS: Questionnaires were returned for 22,161 HIV-negative or status-unknown participants. While 85.3% had ever tested for HIV, 57.6% had tested in the last 12 months. Recent testing was greater in those living in Sydney, in younger men, in gay-identified men, in gay community-attached men, in those who reported unprotected anal intercourse and a higher number of sexual partners, and in partners of HIV-positive men. Although recent testing declined from 1996 to 2001, this trend was no longer significant when adjusted for other testing predictors. CONCLUSIONS: In Australia, HIV testing among gay men decreased slightly from 1996 to 2001, but the trend was not significant when adjusted for other predictors. Testing levels were highest among those at highest risk of HIV infection, and lowest among non gay-identified and non gay-community attached homosexual men.

[Transdermal delivery of cyclosporin A solubilized in mixed micelles through mice skin].

AIM: To investigate the transdermal delivery effects of cyclosporine A solubilized in mixed micelles composed of phospholipid and different surfactants. METHODS: When applied onto the excised abdominal skin of the mice occlusively, the enhancing effects of various mixed micelles on the penetration of cyclosporin A were assessed by an in vitro permeation technique. In vivo study was carried out by topical application of sodium cholate-phospholipid mixed micelles onto the mice skin and drug blood concentration was detected. RESULTS: In vitro, mixed micelles containing different surfactants displayed distinct permeability and corresponded to the following order: sodium cholate > sodium deoxycholate > Trition X-100 > Tween-20. In vivo, peak drug concentration was detected at 5 h and after that the concentration fell down slowly. CONCLUSION: Mixed micelles were shown to be efficient carrier for the transdermal delivery of the lipophilic polypeptide when kept in solution during the application process.

Albumin inhibits apolipoprotein AI and AII production in human hepatoblastoma cell line (Hep G2): additive effects of oleate-albumin complex.

Although the role of multiple humoral agents (such as plasma albumin, glucose, hormones etc.) are implicated in lipoprotein metabolism, the mechanism of action of these agents on various steps of the synthesis and secretion of lipoproteins and apolipoproteins (protein moieties of lipoproteins) are not completely understood. Specifically, the hepatocellular mechanisms of the effect of albumin and fatty acids on apolipoprotein (apo) AI and AII [major proteins of high density lipoproteins (HDL)] synthesis and secretion are not known. Using human hepatoblastoma cells (Hep G2) as an in vitro model system, this study examined the effect of albumin and fatty acids on the synthesis, secretion, and the steady-state mRNA expression of apo AI and AII. The data indicated that the incubation of Hep G2 cells with albumin, dose-dependently, inhibited apo AI and AII accumulation (secretion) in the media, de novo synthesis, and the steady-state mRNA expression. Albumin did not alter total protein synthesis; thus the effect of albumin appeared to be specific for the synthesis and secretion of apo AI and apo AII. Free fatty acids (FFA) are transported by albumin and diseases characterized by enhanced FFA mobilization (e.g. diabetes mellitus) are associated with low HDL levels. Studies were therefore performed to examine the effect of albumin-bound-oleic acid on apo AI and apo AII production. The results showed that the albumin-oleate complex further increased the inhibitory effects of albumin on apo AI and apo AII production. These data suggest how HDL metabolism may be affected at the hepatocellular level by alterations in plasma albumin concentrations and/or fatty acid mobilization in clinical situations characterized by altered HDL levels.

[Effect of processing contents of saccharides in Danggui].

OBJECTIVE: To explore the effect of processing on the contents of saccharides in Danggui. METHOD: The contents of reducing saccharides, water-soluble saccharides and poly saccharides in Danggui and its processed products were determined by phenol-H2SO4 method. RESULTS: The contents of saccharides in the alcohol-broiled product were higher than those in the crude drug. CONCLUSION: Different processing methods have significant effects on the above saccharides, which administers to the interpreparation of the theory of Danggui processing.

Niacin accelerates intracellular ApoB degradation by inhibiting triacylglycerol synthesis in human hepatoblastoma (HepG2) cells.

The mechanism by which the potent drug niacin decreases apoB-containing atherogenic lipoproteins and prevents coronary disease is unclear. Utilizing human hepatoblastoma (HepG2) cells as an in vitro model, we have examined the effect of niacin on intracellular degradation of apoB and the regulatory mechanisms involved in apoB processing. Niacin significantly increased apoB degradation in a dose- and time-dependent manner. Treatment of HepG2 cells with calpain inhibitor I [N-acetyl-leucyl-leucyl-norleucinal (ALLN), an inhibitor of certain protease-mediated apoB degradation], did not alter niacin-induced apoB degradation. Niacin decreased inhibition of oleate-mediated apoB degradation. Niacin dose-dependently inhibited the synthesis of both fatty acids and triacylglycerol (TG) by 20% to 40% as determined by the incorporation of 14C-acetate and 3H-glycerol into fatty acids and TG, respectively. Incubation of HepG2 cells with niacin significantly inhibited (by 12% to 15%) fatty acid esterification to produce TG as assessed by the incorporation of 3H-oleic acid into TG. 14C-acetate incorporation into cholesterol and phospholipids was unchanged. The activity of microsomal triglyceride transfer protein (MTP), a carrier protein for lipids, was not altered by pretreatment of cells with niacin. ApoB mRNA expression and 125I-LDL protein uptake were also unchanged. These data indicate that niacin accelerates hepatic intracellular post-translational degradation of apoB by selectively reducing triglyceride synthesis (through inhibiting both fatty acid synthesis and fatty acid esterification to produce TG) without affecting ALLN-inhibitable protease- or MTP-mediated intracellular apoB processing, resulting in decreased apoB secretion and hence lower circulating levels of the atherogenic lipoproteins.

Estradiol stimulates apolipoprotein A-I- but not A-II-containing particle synthesis and secretion by stimulating mRNA transcription rate in Hep G2 cells.

Estrogen therapy increases plasma HDL levels, which may reduce cardiovascular risk in postmenopausal women. The mechanism of action of estrogen in influencing various steps in hepatic HDL and apolipoprotein (apo) A-I synthesis and secretion are not fully understood. In this study, we have used the human hepatoblastoma cell line (Hep G2) as an in vitro model system to delineate the effect of estradiol on multiple regulatory steps involved in hepatic HDL metabolism. Incubation of Hep G2 cells with estradiol resulted in the following statistically significant findings: (1) increased accumulation of apoA-I in the medium without affecting uptake/removal of radiolabeled HDL-protein; (2) accelerated incorporation of [3H]leucine into apoA-I; (3) selective increase in [3H]leucine incorporation into lipoprotein (LP) A-I but not LP A-I+A-II HDL particles (HDL particles without and with apoA-II, respectively); (4) increased ability of apoA-I-containing particles to efflux cholesterol from fibroblasts; (5) stimulated steady state apoA-I but not apoA-II mRNA expression; and (6) increased newly transcribed apoA-I mRNA message without effect on apoA-I mRNA half-life. The data indicate that estradiol stimulates newly transcribed hepatic apoA-I mRNA, resulting in a selective increase in LP A-I, a subfraction of HDL that is associated with decreased atherosclerotic cardiovascular disease, especially in premenopausal women.

Niacin decreases removal of high-density lipoprotein apolipoprotein A-I but not cholesterol ester by Hep G2 cells. Implication for reverse cholesterol transport.

Niacin (nicotinic acid) is the most potent clinically used agent for increasing plasma HDL and apolipoprotein (apo) A-I. The mechanism by which niacin increases apoA-I is not clearly understood. We have examined the effect of niacin on the hepatic production and removal of apoA-I using Hep G2 cells as an in vitro model. Incubation of Hep G2 cells with niacin resulted in increased accumulation of apoA-I in the medium in a dose-dependent manner. Incorporation of [3H]leucine and [35S]methionine into apoA-I and apoA-I mRNA expression was unchanged by niacin, suggesting that it did not affect apoA-I de novo synthesis. Uptake of radiolabeled HDL protein and HDL apoA-I by Hep G2 cells was significantly reduced to as much as 82.9 +/- 2.2% (P = .04) and 84.2 +/- 2.8% (P = .02), respectively, of the baseline with increasing concentrations of niacin (0 to 3.0 mmol/L). Specific 125I-HDL protein uptake measured with a 50-fold excess of unlabeled HDL was reduced to as much as 78.3 +/- 4.8% (P = .005) in niacin-treated cells. The uptake of labeled cholesterol esters in HDL was unaffected by niacin. Niacin also effected a similar decrease in HDL protein uptake, but not cholesterol esters, from apoA-I-containing HDL particles isolated by immunoaffinity. The conditioned medium obtained from Hep G2 cells incubated with niacin significantly (P = .002) increased cholesterol efflux from cultured human fibroblasts. These data indicate a novel mechanism whereby niacin selectively decreases hepatic removal of HDL apoA-I but not cholesterol esters, thereby increasing the capacity of retained apoA-I to augment reverse cholesterol transport.

Secretory leukocyte protease inhibitor: a macrophage product induced by and antagonistic to bacterial lipopolysaccharide.

To explore regulation of potentially lethal responses to bacterial lipopolysaccharide (LPS), we used differential display under LPS-free conditions to compare macrophage cell lines from two strains of mice congenic for a locus affecting LPS sensitivity. LPS-hyporesponsive cells, primary macrophages, and polymorphonuclear leukocytes transcribed secretory leukocyte protease inhibitor (SLPI), a known epithelial cell-derived inhibitor of leukocyte serine proteases. Transfection of macrophages with SLPI suppressed LPS-induced activation of NF-kappa B and production of nitric oxide and TNF alpha. The ability of interferon-gamma (IFN gamma) to restore LPS responsiveness is a hallmark of the LPS-hyporesponsive phenotype. IFN gamma suppressed expression of SLPI and restored LPS responsiveness to SLPI-producing cells. Thus, SLPI is an LPS-induced IFN gamma-suppressible phagocyte product that serves to inhibit LPS responses.

Effect of serum subfractions from peritoneal dialysis patients on Hep-G2 cell apolipoprotein A-I and B metabolism.

We previously showed that uremic serum subfractions isolated from hemodialysis (HD) patients inhibited the production of apolipoprotein (apo) A-I by human hepatoblastoma cells, Hep-G2. Because of the reported differences in atherogenic cardiovascular mortality between HD and peritoneal dialysis (PD) patients, we examined the effect of similar subfractions from PD patients on apo A-I and apo B synthesis. After obtaining informed consent, serum samples from five normal subjects and nine stable PD patients were applied to Sephadex G-25 columns to obtain the serum subfractions used in the various experiments. Sephadex G-25 chromatograms of PD sera showed a broad peak from fractions 30 through 60 (molecular wt 500 to 2000 Da). Control serum showed no peak in this region. PD serum subfractions decreased apo A-I synthesis, secretion, and apo A-I mRNA expression by Hep-G2 cells when compared to subfractions from control subjects. Cholesterol efflux studies showed that conditioned media obtained from Hep-G2 cells incubated with PD serum subfractions inhibited cholesterol efflux from fibroblasts, suggesting a biologically-significant decrease in apo A-I synthesis. PD serum subfractions increased protein synthesis and mRNA expressions of apo B by Hep-G2 cells. Therefore, serum subfractions obtained from PD patients decreased apo A-I and increased apo B synthesis, findings consistent with their serum lipoprotein profiles suggesting that a biologically-active component in these subfractions could contribute to the risk of atherogenic cardiovascular disease in PD.

Gemfibrozil stimulates apolipoprotein A-I synthesis and secretion by stabilization of mRNA transcripts in human hepatoblastoma cell line (Hep G2).

Gemfibrozil is a widely used drug that elevates plasma HDL and lowers triglycerides and LDL. The mechanism of action of this pharmacological agent on HDL metabolism is not established. Since the liver is the major organ involved in HDL production and removal, we assessed the effect of gemfibrozil on the modulation of apoA-I (a major protein of HDL)-containing particles by a human hepatoblastoma cell line (Hep G2). Incubation of Hep G2 cells with gemfibrozil resulted in the following statistically significant findings: (1) increased accumulation of apoA-I in the medium without affecting uptake of radiolabeled HDL-protein or HDL-apoA-I; (2) accelerated incorporation of [3H]leucine and [35S]methionine into apoA-I; (3) equivalent increases in [3H]leucine incorporation into HDL particles without and with apoA-II (LpA-I and LpA-I+A-II, respectively); (4) equal efflux of fibroblast cholesterol by harvested LpA-I and LpA-I+A-II particles; (5) increased steady state apoA-I mRNA without affecting apoA-I transcription; and (6) increased apoA-I mRNA half-life (2.2-fold). These data indicate that gemfibrozil stabilizes apoA-I mRNA transcripts, resulting in increased translation of functional apoA-I-containing particles capable of effluxing cellular cholesterol, thus defining a major mechanism by which gemfibrozil increases HDL.