Relationship between menopausal hormone therapy and colorectal cancer: a cohort study utilizing the health insurance database in South Korea (HISK)-II.

OBJECTIVE: Many studies have demonstrated that menopausal hormone therapy is associated with a reduced risk for colorectal cancer. This study investigated the relationship between specific hormone therapy regimens and colorectal cancer risk in postmenopausal women in South Korea using national insurance claims data. METHODS: This population-based, retrospective cohort study used insurance data provided by the Health Insurance Review and Assessment Service between 2007 and 2020. The hormone therapy group comprised women ≥40 years of age who underwent hormone therapy for the first time between 2011 and 2014. The control group included women ≥40 years of age who visited medical institutions for menopause-related issues during the same period but did not undergo hormone therapy. RESULTS: After 1:1 propensity score matching, 153,736 women were grouped into either the hormone therapy or nonhormone therapy groups. The incidence of colorectal cancer was 46 and 53 per 100,000 person-years in the nonhormone therapy and hormone therapy groups, respectively. Hormone therapy was associated with an increased risk for colorectal cancer (hazard ratio 1.124 [95% confidence interval 1.002-1.261]). Subgroup analysis, according to hormone therapy type, revealed no significant differences in the risk of colorectal cancer for estrogen plus progestogen or estrogen therapy alone; however, tibolone was associated with an increased risk of colorectal cancer compared to nonhormone therapy (hazard ratio, 1.178 [95% confidence interval, 1.021-1.359]). CONCLUSIONS: This study found an increased risk of colorectal cancer in women receiving hormone therapy, and tibolone was significantly associated with an increased risk of colorectal cancer. However, the magnitude of the increase was small and unlikely to be of clinical significance.

Association between breast diseases and symptomatic uterine fibroids by using South Korean National Health Insurance database.

Both the uterus and breasts have sex hormone dependence, yet there are few studies on the association between breast disease and uterine fibroids (UFs). The purpose of this study was to investigate the incidence of benign breast disease (BBD), carcinoma in situ (CIS), and breast cancer (BC) in women treated for UFs compared to women who were not treated for UFs. This retrospective cohort study used national health insurance data from January 1st, 2011, to December 31st, 2020. We selected women between 20 and 50 years old who (1) were treated for UFs (UF group) or (2) visited medical institutions for personal health screening tests without UFs (control group). We analyzed independent variables such as age, socioeconomic status (SES), region, Charlson comorbidity index (CCI), delivery status, menopausal status, menopausal hormone therapy (MHT), endometriosis, hypertension (HTN), diabetes mellitus (DM), and dyslipidemia based on the first date of uterine myomectomy in the UF group and the first visiting date for health screening in the non-UF group. There were 190,583 and 439,940 participants in the UF and control groups, respectively. Compared with those of the control group, the RRs of BBD, CIS, and BC were increased in the UF group. The hazard ratios (HRs) of BBD, CIS, and BC in the UF group were 1.335 (95% confidence interval (CI) 1.299-1.372), 1.796 (95% CI 1.542-2.092), and 1.3 (95% CI 1.198-1.41), respectively. When we analyzed the risk of BC according to age at inclusion, UFs group had the increased risk of BCs in all age groups in comparison with control group. Women with low SES (HR 0.514, 95% CI 0.36-0.734) and living in rural areas (HR 0.889, 95% CI 0.822-0.962) had a lower risk of BC. Our study showed that women with UFs had a higher risk of BBD, CIS, and BC than those without UFs. This result suggests that women with UFs should be more conscious of BC than those without UFs. Therefore, doctors should consider recommending regular breast self-exams, mammography, or ultrasound for the early detection of BC in women with UFs.

The increased risk of colorectal cancer in the women who underwent hysterectomy from the South Korean National Health Insurance Database.

BACKGROUND: Several population-based studies and observational studies have shown that oophorectomy is associated with an increased risk of colorectal cancer (CRC), and hormone replacement therapy has been associated with a reduction in the risk of colorectal cancer. This study was carried out to investigate whether hysterectomy, which may affect the levels of female hormones, is associated with a risk of cancer of the specific gastrointestinal tract. METHODS: This population-based retrospective cohort study was conducted using insurance data provided by the Health Insurance Review and Assessment Service (HIRA) from January 1, 2007, to December 31, 2020. The hysterectomy group included 40- to 59-year-old women who underwent hysterectomy with uterine leiomyoma or uterine endometriosis from January 1, 2011, to December 31, 2014. The control group included women aged 40 to 59 years who visited medical institutions for medical examination from January 1, 2011 to December 31, 2014. RESULTS: The hysterectomy and non-hysterectomhy groups comprised 66,204 and 89,768 subjects, respectively. The median ages in the non-hysterectomy group and hysterectomy group were 48 (range: 43-53) and 46 (range: 44-49) years, respectively. In the unadjusted results of the analysis, all colorectal cancer (CRC) increased in the hysterectomy alone group (HR 1.222, 95% confidence interval (CI) 1.016-1.47, p = 0.033), sigmoid colon cancer increased in the hysterectomy alone group (HR 1.71, 95% CI 1.073-2.724, p = 0.024), and rectal cancer increased in the hysterectomy with adnexal surgery group (HR 1.924, 95% CI 1.073-2.724, p = 0.002). The adjusted results showed that all CRC increased in the hysterectomy alone group (HR 1.406, 95% CI 1.057-1.871, p = 0.019), colon cancer increased in the hysterectomy alone group (HR 1.523, 95% CI 1.068-2.17, p = 0.02), and rectal cancer increased in the hysterectomy with adnexal surgery group (HR 1.933, 95% CI 1.131-3.302, p = 0.016). The all-cause mortality of GI cancer increased in the hysterectomy alone group (HR 3.495, 95% CI 1.347-9.07, p = 0.001). CONCLUSIONS: This study showed that the risk of all CRC increased in women who underwent hysterectomy compared with women who did not. In particular, the risk of rectal cancer was significantly higher in the women who underwent hysterectomy with adnexal surgery than in the controls. There was no association between hysterectomy and other GI cancers.

Prognostic Factors Affecting Disease-Free Survival and Overall Survival in T4 Colon Cancer.

PURPOSE: It is known that as the T stage of a carcinoma progresses, the prognosis becomes poorer. However, there are few studies about factors that affect the prognosis of T4 advanced colon cancer. This study aimed to identify the prognostic factors associated with disease-free survival (DFS) and overall survival (OS) in T4 colon cancer. METHODS: Patients diagnosed with stage T4 on histopathology after undergoing curative surgery for colon cancer between March 2009 and March 2018 were retrospectively analyzed for factors related to postoperative survival. Primary outcomes were DFS and OS. RESULTS: Eighty-two patients were included in the study. DFS and OS of the pathologic (p) T4b group were not inferior to that of the pT4a group. Multivariate analysis showed that differentiation (hazard ratio [HR], 4.994; P = 0.005), and laparoscopic surgery (HR, 0.323; P = 0.008) were significant prognostic factors for DFS, while differentiation (HR, 7.904; P ≤ 0.001) and chemotherapy (HR, 0.344; P = 0.038) were significant prognostic factors for OS. CONCLUSION: Tumor differentiation, laparoscopic surgery, and adjuvant chemotherapy were found to be significant prognostic factors in patients with T4 colon cancer. Adjuvant chemotherapy and curative resections by laparoscopy might improve the prognosis in these patients.

Prognostic Factors and Management for Left Colonic Perforation: Can Hartmann's Procedure Be Preventable?

PURPOSE: To identify factors significantly associated with the mortality of patients with left colonic perforation, and to compare the outcome of Hartmann's procedure (HP) and primary repair (PR) or primary anastomosis (PA) in patients with left colonic perforation without factors associated with mortality. METHODS: This retrospective study included patients who underwent surgery for left colonic perforation from January 2009 to February 2018. Preoperative factors related to postoperative mortality, including vital signs, laboratory findings, and intraoperative findings, were analyzed by type of operation. The chi-square, Fisher exact, and Mann-Whitney U-tests were used to analyze the data. RESULTS: Ninety-one patients were included (36 men, 55 women), and 15 (16.5%) died postoperatively. Prognostic factors were age, leukopenia, thrombocytopenia, bleeding tendency, acute kidney injury, hemodynamic instability, and the existence of feculent ascites. Leukopenia and longer operative time were independent risk factors for mortality. Seventy-nine patients did not have leukopenia and 30 of these patients who underwent PR without diversion were excluded from the subanalysis. HP was performed in 30 patients, and PR with diversion and PA with or without diversion were performed in 19. Compared to the other operative methods, HP had no advantage in reducing hospital mortality (P=0.458) and morbidity. CONCLUSION: Leukopenia could be an objective prognostic factor for left colonic perforation. Although HP is the gold standard for septic left colonic perforation, it did not improve the hospital mortality of the patients without leukopenia. For such patients, PR or PA may be suggested as an alternative option for left colonic perforation.

Factors Predicting the Need for Early Surgical Intervention for Small Bowel Obstruction.

PURPOSE: Small bowel obstruction (SBO) is a common disease that requires hospitalization. The most common cause of SBO is postoperative adhesion. Delayed timing of operations in patients who need surgical intervention results in mortality or morbidity. A number of studies on SBO have established criteria for emergency surgery. However, few objective clinical parameters are available for screening patients who need a delayed operation. Therefore, we analyzed factors that affect the clinical course of SBO to select appropriate therapeutic plans for reducing the risk of complications in these patients. METHODS: We investigated the clinical characteristics of patients admitted to the surgery department of our hospital between January 1, 2015, and December 31, 2016, who were diagnosed with SBO. Patients were divided into an operative treatment group (n = 12) and a conservative treatment group (n = 96). We compared clinical characteristics between the 2 groups. RESULTS: The operative treatment group underwent more operations before SBO than the conservative treatment group (P = 0.007). Initial leukocyte counts (P = 0.004) and C-reactive protein (CRP) levels (P = 0.028) were elevated in the operative group. Body mass index (BMI) was lower in the operative group (P = 0.013). CONCLUSION: The number of operations before SBO, leukocyte counts, CRP levels, and BMI were useful parameters for selecting patients who needed an urgent operation for SBO.

Unmodifiable Clinicopathological Risk Factors of Shoulder Tip or Subcostal Pain after Laparoscopic Appendectomy.

Purpose: Appendectomy, which comprises most benign intra-abdominal surgeries, is currently assisted by laparoscopy in most cases. However, many patients complain of postoperative shoulder or subcostal pain after laparoscopic surgery. In some cases, the pain lasts even several weeks after surgery. This study aimed to analyze unmodifiable clinicopathological factors of patients who underwent laparoscopic appendectomy and to minimize preoperative and postoperative discomfort. Methods: Patients admitted for laparoscopic appendectomy for acute appendicitis with an American Society of Anesthesiology (ASA) grades I and II, and ages 12~70 years were enrolled in the study. Postoperative shoulder or subcostal pain was assessed using the visual analogue scale (VAS) for pain and analyzed with the clinicopathological factors of the patients, including age, sex, weight, height, body mass index (BMI), and abdominal circumference (AC) difference. Results: Of the 124 patients, 40 complained of postoperative shoulder or subcostal pain with a VAS score of ≥4. The risk of the postoperative shoulder or subcostal pain increased in women (p=0.001). From a univariate analysis, the risk of postoperative shoulder or subcostal pain increased with lower height, weight and BMI (p=0.002, p=0.001, p=0.012) and with greater AC difference (p=0.012). However, a multivariate analysis showed that lower weight was the only risk factor of postoperative pain (p=0.005). Conclusion: The risk of postoperative shoulder or subcostal pain after laparoscopic appendectomy was significantly increased with lower weight.

Factors affecting the negative predictive value of positron emission tomography/computed tomography for axillary lymph node staging in breast cancer patients.

BACKGROUND/OBJECTIVE: Axillary lymph node staging (ALNS) is an important step in the treatment of breast cancer and sentinel lymph node biopsy (SLNB) is a standard procedure for ALNS. Recently, the use of positron emission tomography/computed tomography (PET/CT) for whole body staging in patients with breast cancer has been increasing. The negative predictive value (NPV) of the specific diagnostic modality is the crucial value for excluding axillary lymph node metastasis (ALNM) and guiding the decision not to proceed with axillary lymph node dissection. The aim of this study was to identify patient groups in which PET/CT yields a high NPV for ALNS. METHODS: We reviewed data from the records of 262 patients with breast cancer who underwent PET/CT before surgery between February 2009 and March 2018. We searched for factors associated with pathological axillary lymph node metastasis in patients with negative ALNM on PET/CT. Then, we calculated the NPV of PET/CT for ALNS in the patient group without the identified factors and in all patients. RESULTS: Age ≥75 years and tumor size on ultrasonography (US) ≥ 15 mm were the associated factors; the NPV of PET/CT in patients without these factors compared to all patients was 97.2% versus 88.7%. CONCLUSION: The NPV of PET/CT for ALNS in patients younger than 75 years and with tumor size on US < 15 mm is higher than that in all patients and comparable to the NPV of SLNB reported in previous studies (90.1-97.0%).

Low-level laser therapy with 850 nm recovers salivary function via membrane redistribution of aquaporin 5 by reducing intracellular Ca2+ overload and ER stress during hyperglycemia.

The overall goal is to study the effect of low-level laser therapy (LLLT) on membrane distribution of major water channel protein aquaporin 5 (AQP5) in salivary gland during hyperglycemia. Par C10 cells treated with high glucose (50 mM) showed a reduced membrane distribution of AQP5. The functional expression of AQP5 was downregulated due to intracellular Ca2+ overload and ER stress. This reduction in AQP5 expression impairs water permeability and therefore results in hypo-salivation. A reduced salivary flow was also observed in streptozotocin (STZ)-induced diabetic mice model and the expression of AQP5 and phospho-AQP5 was downregulated. Low-level laser treatment with 850 nm (30 mW, 10 min = 18 J/cm2) reduced ER stress and recovered AQP5 membrane distribution via serine phosphorylation in the cells. In the STZ-induced diabetic mouse, LLLT with 850 nm (60 J/cm2) increased salivary flow and upregulated of AQP5 and p-AQP5. ER stress was also reduced via downregulation of caspase 12 and CHOP. In silico analysis confirmed that the serine 156 is one of the most favorable phosphorylation sites of AQP5 and may contribute to the stability of the protein. Therefore, this study suggests high glucose inhibits phosphorylation-dependent AQP5 membrane distribution. High glucose induces intracellular Ca2+ overload and ER stress that disrupt AQP5 functional expression. Low-level laser therapy with 850 nm improves salivary function by increasing AQP5 membrane distribution in hyperglycemia-induced hyposalivation.

Perioperative Serum Carcinoembryonic Antigen Ratio Is a Prognostic Indicator in Patients With Stage II Colorectal Cancer.

PURPOSE: The aim of this study was to evaluate whether the perioperative carcinoembryonic antigen (CEA) ratio could be used as a determinant for adjuvant therapy after curative surgery in stage II colorectal cancer. METHODS: Data for 119 patients with stage II colorectal cancer who underwent radical surgery between 2010 and 2013 were collected. The perioperative CEA ratio was defined as the postoperative/preoperative serum CEA level, and the patients were grouped according to their perioperative CEA ratios: high ratio (≥0.5) and low ratio (<0.5). Overall survival rates were calculated, and their prognostic significances were analyzed. RESULTS: The overall survival rates of the high and the low perioperative CEA groups were 68.2% and 86.8%, respectively (P = 0.033). In patients with normal preoperative CEA levels (<5 ng/mL), the high perioperative CEA ratio group showed a worse survival rate than the low perioperative CEA ratio group (71.7% vs. 100.0%, P = 0.007). In patients with high preoperative CEA levels (≥5 ng/mL), the high perioperative CEA ratio group showed a worse survival rate than the low perioperative CEA ratio group (33.3% vs. 75.0%, P = 0.036). In the multivariate analysis, perioperative CEA ratio (P = 0.046), age (P = 0.034), and venous invasion (P = 0.015) were independent prognostic factors for survival. CONCLUSION: The perioperative CEA ratio is a prognostic indicator for stage II colorectal cancer. Patients with normal preoperative serum CEA levels might also be considered for adjuvant therapy if their perioperative CEA ratios are higher than 0.5.

Inhibition of senescence and promotion of the proliferation of chondrocytes from articular cartilage by CsA and FK506 involves inhibition of p38MAPK.

Cyclosporine A (CsA) and tacrolimus (FK506) are the most important immunosuppressive compounds that block the activation of helper T-cells. In this study, we investigated the effects of CsA and FK506 on growth and senescence of articular chondrocytes. Chondrocytes from young rabbit cartilage entered senescence after 8.6 ± 0.8 population doublings (PDs), while chondrocytes treated with CsA and FK506 entered senescence after 12.3 ± 1.4 and 13.7 ± 0.6 PDs, respectively. Furthermore, chondrocytes from the cartilage of old rabbits were senescent after 2.6 ± 0.9 PDs, whereas those treated with CsA and FK506 were senescent after 8.2 ± 1.8 and 6.9 ± 1.6 PDs, respectively. These compounds also inhibited senescence induction of chondrocytes in a high-cell density pellet culture system. We previously reported that p38MAPK plays a critical role in the onset of senescence in chondrocyte. This study revealed that the phosphorylation of p38MAPK was inhibited by either CsA or FK506. The early onset of senescence in chondrocyte harboring MKK6E, which is a constitutively-active form of MKK6 and increases p38MAPK phosphorylation, was blocked by CsA. These results suggest that CsA and FK506 increase the proliferation and inhibit the senescence of articular chondrocytes through inactivation of p38MAPK.

Identification of a novel anti-apoptotic E3 ubiquitin ligase that ubiquitinates antagonists of inhibitor of apoptosis proteins SMAC, HtrA2, and ARTS.

Identification of new anti-apoptotic genes is important for understanding the molecular mechanisms underlying apoptosis and tumorigenesis. The present study identified a novel anti-apoptotic gene named AREL1, which encodes a HECT (homologous to E6-AP carboxyl terminus) family E3 ubiquitin ligase. AREL1 interacted with and ubiquitinated IAP antagonists such as SMAC, HtrA2, and ARTS. However, AREL1 was cytosolic and did not localize to nuclei or mitochondria. The interactions between AREL1 and the IAP antagonists were specific for apoptosis-stimulated cells, in which the IAP antagonists were released into the cytosol from mitochondria. Furthermore, the ubiquitination and degradation of SMAC, HtrA2, and ARTS were significantly enhanced in AREL1-expressing cells following apoptotic stimulation, indicating that AREL1 binds to and ubiquitinates cytosolic but not mitochondria-associated forms of IAP antagonists. Furthermore, the anti-apoptotic role of AREL1-mediated degradation of SMAC, HtrA2, and ARTS was shown by simultaneous knockdown of three IAP antagonists, which caused the inhibition of caspase-3 cleavage, XIAP degradation, and induction of apoptosis. Therefore, the present study suggests that AREL1-mediated ubiquitination and degradation of cytosolic forms of three IAP antagonists plays an important role in the regulation of apoptosis.

DNA damage induces the IL-6/STAT3 signaling pathway, which has anti-senescence and growth-promoting functions in human tumors.

IL-6 is a multifunctional cytokine that is important for immune responses, cell survival, apoptosis, and proliferation. However, little is known about the correlation between the IL-6 signaling pathway and DNA damage in human tumors. The present study demonstrates the role of the IL-6/STAT3 signaling pathway in human tumor cells exposed to DNA damage. Tumor cells exposed to DNA damage increase the expression and secretion of IL-6 and the phosphorylation of JAK1 and STAT3. The activation of the JAK1-STAT3 signaling pathway is inhibited by knockdown of gp130 or neutralization of soluble IL-6, implying that DNA damage induces the phosphorylation of JAK1 and STAT3 by autocrine IL-6. Interestingly, inhibition of the IL-6/STAT3 signaling pathway impairs the growth of tumor cells exposed to DNA damage and results in the induction of senescence. Therefore, the present study suggests that IL-6 inhibits senescence but promotes the survival and proliferation of tumor cells exposed to DNA damage through the activation of the JAK1-STAT3 signaling pathway.

NF-Y binds to both G1- and G2-specific cyclin promoters; a possible role in linking CDK2/Cyclin A to CDK1/Cyclin B.

We previously reported that CDK2/Cyclin A can phosphorylate and activate the transcription factor NF-Y. In this study, we investigated a potential regulatory role for NF-Y in the transcription of Cyclin A and other cell cycle regulatory genes. Gel-shift assays demonstrate that NF-Y binds to CCAAT sequences in the Cyclin A promoter, as well as to those in the promoters of cell cycle G2 regulators such as CDC2, Cyclin B and CDC25C. Furthermore, expression of Cyclin A increases NF-Y's affinity for CCAAT sequences in the CDC2 promoter; however, Cyclin A's induction of CDC2 transcription is antagonized by p21, an inhibitor of CDK2/Cyclin A. These results suggest a model wherein NF-Y binds to and activates transcription from the Cyclin A promoter, increasing cellular levels of Cyclin A/CDK2 and potentiating NF-Y's capacity for transcriptional transactivation, and imply a positive feedback loop between NF-Y and Cyclin A/CDK2. Our findings are additionally indicative of a role for Cyclin A in activating Cyclin B/CDK1 through promoting NF-Y dependent transcription of Cyclin B and CDC2; NF-Y mediated crosstalk may therefore help to orchestrate cell-cycle progression.

Interferon-gamma-induced expressions of heat shock protein 60 and heat shock protein 10 in C6 astroglioma cells: identification of the signal transducers and activators of transcription 3-binding site in bidirectional promoter.

Heat shock protein 60 and heat shock protein 10 are mitochondrial chaperonin proteins. Here, we report that the expressions of heat shock protein 60 and heat shock protein 10 were upregulated in interferon-gamma-treated C6 astroglioma cells, and the 582 bp in the bidirectional promoter of the heat shock protein 60 and heat shock protein 10 genes is responsible for interferon-gamma-induced induction. The induction of heat shock protein 60 and heat shock protein 10 by interferon-gamma was virtually abolished by introducing mutations into the putative signal transducers and activators of transcription 3-response element in the promoter, and the same mutation impaired increment of the signal transducers and activators of transcription 3-binding after interferon-gamma treatment. Moreover, Rac1 GTPase was required for maximal heat shock protein 10 and heat shock protein 60 inductions by interferon-gamma. These results suggest that interferon-gamma-induced upregulations of heat shock protein 60 and heat shock protein 10 in C6 astroglioma cells are mediated by the signal transducers and activators of transcription 3-binding site, localized in the bidirectional promoter.

Ethyl pyruvate attenuates kainic acid-induced neuronal cell death in the mouse hippocampus.

Recent studies have shown that ethyl pyruvate (EP) acts as an anti-inflammatory molecule in several cell lines including RAW264.7 macrophages. However, the potential therapeutic value of EP for the treatment of the pathologic brain has not been investigated fully. In the present study, we examined whether EP has a beneficial effect on KA-induced neuronal cell death. Intracerebroventricular (i.c.v.) injection of 0.94 nmol (0.2 mug) of KA produced typical neuronal cell death in the CA1 and CA3 pyramidal layers of the hippocampus, and the systemic administration of EP significantly attenuated KA-induced neuronal cell death in these regions. Ethyl pyruvate was found to exert a protective effect when it was injected as late as 12 hr after KA-injection. Moreover, this EP-induced neuroprotection was accompanied by reduced levels of reactive gliosis and COX-2, IL-1beta, and TNF-alpha in the hippocampus. In addition, in passive avoidance tests, KA-induced memory impairment was improved markedly by EP. These results suggest that EP has a therapeutic potential for suppressing KA-induced pathogenesis in the brain.

Enhanced transfection of primary cortical cultures using arginine-grafted PAMAM dendrimer, PAMAM-Arg.

PAMAM-Arg is a cationic arginine-grafted polyamidoamine (PAMAM) dendrimer. In the previous study, we reported that PAMAM-Arg facilitates transfection in a range of mammalian cell types. In the present study, we investigated the transfection efficiency of PAMAM-Arg in primary cortical cultures, which are known to be extremely vulnerable to exogenous gene transfection. PAMAM-Arg/DNA complexes showed particularly high transfection efficiencies and low cytotoxicity in primary cortical cells, as compared to other gene carriers such as, native PAMAM, polyethylenimine (BPEI), and Lipofectamine. Efficient transfection was not limited to neurons but extended to all three glial cells, astrocytes, microglia, and oligodendrocytes, present in these primary cortical cultures. The potential use of PAMAM-Arg was demonstrated by efficient gene knock-down by transfecting HMGB1 shRNA-expressing plasmid. The numbers of green fluorescent protein (GFP)-positive and HMGB1-negative cells indicated that PAMAM-Arg/shRNA-expressing plasmid complex suppressed target gene expression in over 40% of cells, which is the highest level achieved to date in primary cortical culture by any gene carrier. Here, we present evidence of the successful delivery and expression of both a reporter gene and of a shRNA-expressing plasmid in primary cortical cells, which demonstrates the potential of PAMAM-Arg for mediating gene delivery to primary neuronal cells.

Enhanced transfection efficiency of PAMAM dendrimer by surface modification with L-arginine.

We designed a novel type of arginine-rich dendrimer, with a structure based on the well-defined dendrimer, polyamidoamine dendrimer (PAMAM). Further characterization was performed to prove that the polymer is a potent nonviral gene delivery carrier. The primary amines located on the surface of PAMAM were conjugated with L-arginine to generate an L-arginine-grafted-PAMAM dendrimer (PAMAM-Arg). For comparison, an L-lysine-grafted-PAMAM dendrimer (PAMAM-Lys) was also generated and compared as a control reagent. The polymers were found to self-assemble electrostatically with plasmid DNA, forming nanometer-scale complexes. From dynamic light scattering experiments, the mean diameter of the polyplexes was observed to be around 200 nm. We used PicoGreen reagent as an efficient probe for assaying complex formation of polymers with plasmid DNA. The complex composed of PAMAM-Arg/DNA showed increased gene delivery potency compared to native PAMAM dendrimer and PAMAM-Lys. The cytotoxicity and transfection efficiencies for 293, HepG2, and Neuro 2A cells were measured by comparison with PEI and PAMAM. In addition, transfection experiments were performed in primary rat vascular smooth muscle cells, and PAMAM-Arg showed much enhanced transfection efficiency. These findings suggest that the L-arginine-grafted-PAMAM dendrimer possesses the potential to be a novel gene delivery carrier for gene therapy.

Administration of the p38 MAPK inhibitor SB203580 affords brain protection with a wide therapeutic window against focal ischemic insult.

We have reported previously the delayed and differential induction of p38alpha and p38beta mitogen-activated protein kinases (MAPKs) in microglia and astrocytes, respectively, in brain after transient global ischemia. We report here the sustained induction and activation of p38alpha MAPK in activating microglia in rat brain after transient middle cerebral artery occlusion (MCAO). The intraventricular administration of SB203580, a p38 MAPK inhibitor, 30 min before MCAO reduced the infarct volume to 50% of the control, which was accompanied by the significant improvement of neurological deficits. More interestingly, the infarct volume was reduced to 72% and 77% when SB203580 was administered 6 hr and 12 hr after MCAO, respectively. The induction of various factors involved in inflammatory processes, such as inducible nitric oxide synthase (iNOS), tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta) and cyclooxygenase-2 (COX-2), was suppressed by the administration of SB203580 at 6 hr after MCAO. These results suggest that sustained activation of p38 MAPK pathway and p38 MAPK-associated inflammatory processes play a crucial role in postischemic brain.