Regression of Congenital Melanocytic Nevus with Halo Phenomenon Induced by Serial Excision / 대한피부과학회지

Congenital melanocytic nevus is a hamartoma derived from the neural crest that is present at birth. Regression following surgical excision with an apparent halo phenomenon through suture lines has never been reported. A nine-year-old boy presented with a solitary symmetric, oval-shaped, blackish pigmented patch on his right forearm.He reported increasing size of the lesion with no other subjective symptoms. Histological examination of the first excisional biopsy revealed congenital melanocytic nevus, and serial excisions were planned. Interestingly, at the second visit at 18 months after the first biopsy, the size of the congenital melanocytic nevus was reduced with a peripheral whitish halo. Linear regression through suture lines and a peripheral halo was observed after the second and third serial excisions. The mechanism of the halo phenomenon remains elusive but is suggested theorized to be caused by destruction of melanocytes by immune responses of autoantibodies or cytotoxic T cells.

The Efficacy of Topical Tacrolimus Ointment on Cheilitis Induced by Isotretinoin Treatment in Acne Vulgaris Patients / 대한피부과학회지

Background@#Isotretinoin is a gold standard treatment for moderate to severe acne vulgaris but is associated with cheilitis. Though moisturizing agents or petrolatum have been suggested, uncomfortable isotretinoin-induced cheilitis can disrupt treatment of acne patients. Topical tacrolimus has been used in atopic dermatitis with good safety and efficacy; however, there is no study of application of topical tacrolimus in cheilitis induced by isotretinoin. @*Objective@#In this study, we aimed to describe the efficacy of topical tacrolimus 0.1% ointment on cheilitis associated with isotretinoin therapy using isotretinoin cheilitis grading scale (ICGS), investigator global assessment (IGA), and patient global assessment (PGA). We also observed the side effects of topical tacrolimus ointment. @*Methods@#Fifty acne vulgaris patients with isotretinoin-induced cheilitis were randomly allocated to either topical tacrolimus or petrolatum treatment using permuted-block randomization. Patients were followed-up at 4 and 8 weeks, at which cheilitis lesions were photographed and evaluated with ICGS, IGA, and PGA. @*Results@#Compared to petrolatum group, tacrolimus group had greater responses to treatment as measured by mean values of ICGS, IGA, PGA at follow-up visits. Also, the ICGS decrease was larger in the tacrolimus group compared with the petrolatum group even according to isotretinoin dose. @*Conclusion@#Topical tacrolimus ointment had superior efficacy in treating cheilitis induced by isotretinoin compared to petrolatum. Erythema, fissures, scales, and commissures all showed better response to tacrolimus ointment. Topical tacrolimus ointment can be administered as an effective strategy in treatment of cheilitis as a complication of isotretinoin therapy and can improve compliance of acne patients.

Regression of Congenital Melanocytic Nevus with Halo Phenomenon Induced by Serial Excision / 대한피부과학회지

Congenital melanocytic nevus is a hamartoma derived from the neural crest that is present at birth. Regression following surgical excision with an apparent halo phenomenon through suture lines has never been reported. A nine-year-old boy presented with a solitary symmetric, oval-shaped, blackish pigmented patch on his right forearm.He reported increasing size of the lesion with no other subjective symptoms. Histological examination of the first excisional biopsy revealed congenital melanocytic nevus, and serial excisions were planned. Interestingly, at the second visit at 18 months after the first biopsy, the size of the congenital melanocytic nevus was reduced with a peripheral whitish halo. Linear regression through suture lines and a peripheral halo was observed after the second and third serial excisions. The mechanism of the halo phenomenon remains elusive but is suggested theorized to be caused by destruction of melanocytes by immune responses of autoantibodies or cytotoxic T cells.

The Efficacy of Topical Tacrolimus Ointment on Cheilitis Induced by Isotretinoin Treatment in Acne Vulgaris Patients / 대한피부과학회지

Background@#Isotretinoin is a gold standard treatment for moderate to severe acne vulgaris but is associated with cheilitis. Though moisturizing agents or petrolatum have been suggested, uncomfortable isotretinoin-induced cheilitis can disrupt treatment of acne patients. Topical tacrolimus has been used in atopic dermatitis with good safety and efficacy; however, there is no study of application of topical tacrolimus in cheilitis induced by isotretinoin. @*Objective@#In this study, we aimed to describe the efficacy of topical tacrolimus 0.1% ointment on cheilitis associated with isotretinoin therapy using isotretinoin cheilitis grading scale (ICGS), investigator global assessment (IGA), and patient global assessment (PGA). We also observed the side effects of topical tacrolimus ointment. @*Methods@#Fifty acne vulgaris patients with isotretinoin-induced cheilitis were randomly allocated to either topical tacrolimus or petrolatum treatment using permuted-block randomization. Patients were followed-up at 4 and 8 weeks, at which cheilitis lesions were photographed and evaluated with ICGS, IGA, and PGA. @*Results@#Compared to petrolatum group, tacrolimus group had greater responses to treatment as measured by mean values of ICGS, IGA, PGA at follow-up visits. Also, the ICGS decrease was larger in the tacrolimus group compared with the petrolatum group even according to isotretinoin dose. @*Conclusion@#Topical tacrolimus ointment had superior efficacy in treating cheilitis induced by isotretinoin compared to petrolatum. Erythema, fissures, scales, and commissures all showed better response to tacrolimus ointment. Topical tacrolimus ointment can be administered as an effective strategy in treatment of cheilitis as a complication of isotretinoin therapy and can improve compliance of acne patients.

Effect of Epidermal Growth Factor-Containing Ointment for the Management of Laser-Induced Postinflammatory Erythema and Hyperpigmentation / 대한피부과학회지

Background@#Epidermal growth factor (EGF) stimulates wound healing in various skin conditions, and EGFcontaining formulations can promote wound regeneration and help reduce complications after laser treatment. @*Objective@#To evaluate the effect of EGF-containing ointment on wound regeneration after ablative laser treatment and laser-induced postinflammatory erythema (PIE) and hyperpigmentation (PIH). @*Methods@#Twenty-five patients who required treatment with ablative laser on both sides of the face were enrolled and underwent one session of laser treatment. Postoperatively, all patients were instructed to apply the EGFcontaining ointment on the randomly assigned-hemifacial side, while gentamicin cream was applied on the opposite side. Investigators’ global assessment (IGA), evaluating general wound condition, erythema, pigmentation, and scab shedding, and patient’s global assessment (PGA), evaluating patient satisfaction with clinical improvement, were both measured at 1 week and 4 weeks after treatment. Objective assessments, including melanin index (MI), erythema index (EI), skin hydration, and transepidermal water loss (TEWL), were measured before treatment and at 1 week and 4 weeks after treatment. @*Results@#The EGF-containing ointment showed better outcomes with IGA (1 week; p=0.02, 4 weeks; p=0.002) and PGA (1 week; p=0.04, 4 weeks; p=0.01) scores compared to gentamicin cream. Compared to gentamicin cream, treatment with EGF-containing ointment resulted in significantly lower EI (p=0.044) and MI (p=0.023), but skin hydration and TEWL between the two groups exhibited nonsignificant differences. @*Conclusion@#EGF-containing ointment could be an effective adjuvant option for wound regeneration after ablative laser treatment and minimizing laser-induced PIE and PIH in Asian patients.

A Case of Livedoid Vasculopathy Successfully Treated with Sulodexide

We report a 29-year-old female with a one-month history of non-healing multiple erythematous to violaceous plaques with crusts over both legs and feet. Tender, scarring ulcers with surrounding erythema were present. The clinical manifestation, together with histopathologic findings of fibrinoid plugs within vascular lumens and walls, as well as red blood cell extravasation, led to diagnosis of livedoid vasculopathy.The patient experienced recurrent painful violaceous plaques with ulcerations during the two years of treatment with oral pentoxifylline 400 mg three times daily. The cutaneous lesions and symptoms dramatically improved after the treatment regimen changed to oral sulodexide (250 lipasemic units) three times daily. Sulodexide, a highly purified mixture of glycosaminoglycans including dermatan sulfate and lowmolecular weight heparin, could be an effective therapy for recalcitrant livedoid vasculopathy. Herein, we report a case of livedoid vasculopathy treated with sulodexide, which has not previously been reported.

Innate Lymphoid Cells and Infection

The functions of innate lymphoid cells (ILCs) have been known to play an important role in immunity and immune responses. ILCs are rapidly-responding cells that are involved in tissue remodeling, cancer, the regulation of autoimmune inflammation and resistance to pathogens. Understanding the role of ILCs in regulating immune response could be useful for the development of new therapeutic strategies against emerging or re-emerging infectious diseases. However, the relevance of ILCs in infectious diseases was not fully uncovered. This review provides an overview of the current knowledge of the functional characteristics of ILCs and how these cells interact with pathogens to mediate immune responses.

Characterization of Endoplasmic Reticulum Stress and Apoptosis in Macrophages Infected with Mycobacterium tuberculosis Isolates from Korea Patients

Apoptosis is an important host defense mechanism against mycobacterial infection. Recent reports suggest that links between apoptosis and endoplasmic reticulum (ER) stress are critical for the regulation of mycobacterial survival; however, the exact regulatory mechanisms are not well known. In this study, we isolated 20 Mycobacterium tuberculosis (Mtb) clinical strains from Korean patients and examined ER stress-mediated apoptosis in Mtb-infected macrophages. Most Mtb strains increased the rates of apoptosis and production of ER stress-sensing molecules in mouse macrophages, similar to Mtb H37Rv infection. Moreover, the intracellular survival of Mtb clinical isolates in macrophages was similar to that of H37Rv. Our data suggest that infection with Mtb downregulated MCP-1 and MCPIP. The regulation of MCPIP may decrease ROS production, leading to a reduction in ER stress-mediated apoptosis.

Intrathecal Administration of Mesenchymal Stem Cells Reduces the Reactive Oxygen Species and Pain Behavior in Neuropathic Rats

BACKGROUND: Neuropathic pain induced by spinal or peripheral nerve injury is very resistant to common pain killers, nerve block, and other pain management approaches. Recently, several studies using stem cells suggested a new way to control the neuropatic pain. In this study, we used the spinal nerve L5 ligation (SNL) model to investigate whether intrathecal rat mesenchymal stem cells (rMSCs) were able to decrease pain behavior, as well as the relationship between rMSCs and reactive oxygen species (ROS). METHODS: Neuropathic pain of the left hind paw was induced by unilateral SNL in Sprague-Dawley rats (n = 10 in each group). Mechanical sensitivity was assessed using Von Frey filaments at 3, 7, 10, 12, 14, 17, and 24 days post-ligation. rMSCs (10 microl, 1 x 105) or phosphate buffer saline (PBS, 10 microl) was injected intrathecally at 7 days post-ligation. Dihydroethidium (DHE), an oxidative fluorescent dye, was used to detect ROS at 24 days post-ligation. RESULTS: Tight ligation of the L5 spinal nerve induced allodynia in the left hind paw after 3 days post-ligation. ROS expression was increased significantly (P < 0.05) in spinal dorsal horn of L5. Intrathecal rMSCs significantly (P < 0.01) alleviated the allodynia at 10 days after intrathecal injection (17 days post-ligation). Intrathecal rMSCs administration significantly (P < 0.05) reduced ROS expression in the spinal dorsal horn. CONCLUSIONS: These results suggest that rMSCs may modulate neuropathic pain generation through ROS expression after spinal nerve ligation.

Macrophage Polarization and Infection

Monocytes and macrophages regulate host immune system against infectious pathogens. Activated macrophages play an important role in restricting the multiplication and dissemination of pathogens. The concept of alternative activation of macrophages might provide useful insights into pathology of infectious diseases. M1 macrophages (classically activated macrophages) and M2 macrophages (alternatively activated macrophages) are associated with responses to tissue remodeling, pro-inflammatory and anti-inflammatory reactions in various infectious diseases. However, the relevance of macrophage polarization in several infectious diseases was not revealed clearly. Macrophage plasticity and polarization should be considered as a useful conceptual framework for understanding the unknown pathogenesis of infectious diseases. Here we reviewed the recent progress on macrophage polarization and its characters in infectious diseases.

Cell Death and Bacterial Infection

Cells can die through various biochemical pathways related to complex pathophysiological process. Different types of cell death are closely associated with microbial infection. Several regulatory mechanisms of cell death during bacterial infection play important roles to control the pathogens. Bacteria usually manipulate host defense mechanisms to survive and eventually replicate. Host cell death is one of the intrinsic immune defense mechanisms even if infected cells were sacrificed and it induced detrimental effects on host. Understanding the role of cell death during bacterial infection is important to provide insight into the pathogenesis of unknown infectious diseases. In this review, the different forms of cell death are discussed.

Endoplasmic Reticulum Stress Responses and Apoptosis

The endoplasmic reticulum (ER) plays a crucial role in various cellular activities and cell survival. Almost all of the resident proteins usually enter the ER, and are modified with N-linked glycans and folded into the appropriate secondary and tertiary structures. When cells are faced with stressful conditions, unfolded proteins are accumulated in the ER. The discrepancies between the protein folding capacities and client protein load lead to ER stress. If the stress is prolonged, ER stress responses can activate apoptosis. ER stress-mediated apoptosis is implicated in the pathophysiology of human diseases, including several neurodegenerative diseases, diabetes mellitus, and various infectious diseases. Thus, the ER is now considered as an important organelle that can decide cell survival or death. In this review, the recent progress on ER stress and apoptosis is summarized.

Phenotypic and Genotypic Correction of WASP Gene Mutation in Wiskott-Aldrich Syndrome by Unrelated Cord Blood Stem Cell Transplantation

We present two cases of Wiskott-Aldrich syndrome (WAS), in which nonsense mutations in the WASP gene were corrected phenotypically as well as genotypically by unrelated cord blood stem cell transplantation (CBSCT). Two male patients were diagnosed with WAS at the age of 5-month and 3-month and each received unrelated CBSCT at 16-month and 20-month of age, respectively. The infused cord blood (CB) units had 4/6 and 5/6 HLA matches and the infusion doses of total nucleated cells (TNC) and CD34+ cells were 6.24x10(7)/kg and 5.08x10(7)/kg for TNC and 1.33x10(5)/kg and 4.8x10(5)/kg for CD34+ cells, for UPN1 and UPN2, respectively. Complete donor cell chimerism was documented by variable number tandem repeat (VNTR) with neutrophil engraftment on days 31 and 13 and platelets on days 58 and 50, respectively. Immunologic reconstitution demonstrated that CBSCT resulted in consistent and stable T-, B-, and NK-cell development. Flow cytometric analysis for immunologic markers and sequence analysis of the WASP gene mutation revealed a normal pattern after CBSCT. These cases demonstrate that CBs can be an important source of stem cells for the phenotypical and genotypical correction of genetic diseases such as WAS.

Differential Roles of Toll-like Receptor (TLR) 2 and 4 between PPD- and 38-kDa-induced Proinflammatory Cytokine Productions in Human Monocytes

In this study, we investigated the role of toll-like receptor (TLR) and mitogen-activated protein kinase (MAPK) pathways involved in the tumor necrosis factor (TNF)-alpha and interleukin (IL)-6 expression after stimulation with purified protein derivatives (PPD) or native 38-kDa protein antigen (Ag) of Mycobacterium tuberculosis H37Rv in human primary monocytes. Both PPD and 38-kDa Ag significantly induced TNF-alpha and IL-6 in human primary monocytes. MAPK [extracellular signal-regulated kinase (ERK) 1/2 and p38] are rapidly phosphorylated in human monocytes stimulated with the PPD or 38-kDa Ag. Both p38 and ERK 1/2 activation are essential for PPD- or 38-kDa-induced TNF-alpha and IL-6 production. The inhibition of TLR2 and TLR4 by specific antibodies significantly abrogated the 38-kDa-induced secretion of TNF-alpha and IL-6, whereas blockade of TLR2, but not TLR4, was responsible for the PPD-induced TNF-alpha and IL-6 production in human monocytes. Collectively, these data suggest that the PPD and 38-kDa Ag differentially interact with TLR2 and TLR4, which in turn mediate an essential role for the early inflammatory immune responses during human tuberculosis.

Identification and Functional analysis of Gene Expression in Mycobacterium tuberculosis-infected Human Monocytic Cells Under Hypoxic Conditions

Mycobacterium tuberculosis-induced granulomatous lesions, particularly those undergoing central caseation, are known as hypoxic. To analyze the host genes associated with hypoxic conditions from cells infected with M. tuberculosis, we performed GeneChip analyses on mRNA from M. tuberculosis H37Rv-treated human monocytic THP-1 cells cultured in oxygen-depleted status for 18 h. The expression of 99 genes was altered, including those involved in intracellular signaling, energy production, and protein metabolism, as revealed by stringent microarray data analysis. Most notably, mRNA expression of chemokine macrophage inflammatory protein 3alpha/CC chemokine ligand 20 (CCL20) was significantly up-regulated in M. tuberculosis-infected cells under hypoxic conditions. We further analyzed the CCL20 expression in peripheral blood mononuclear cells (PBMCs) and monocyte derived macrophages (MDMs) from healthy controls and TB patients. A comparative analysis has revealed that the mRNA and protein expression of CCL20 were prominently up-regulated in PBMCs, and MDMs from TB patients, compared with healthy controls. Collectively, these data show that the gene expression of CCL20 was up-regulated in M. tuberculosis H37Rv-infected human monocytic THP-1 cells cultured in hypoxic conditions. In addition, the production of CCL20 is substantially increased in cells from TB patients than in healthy controls, suggesting an important role of CCL20 in the immunopathogenesis during TB infection.

Purification of Native Ag85 Complex, 38-kDa and MTB12 Protein Antigens from the Culture Filtrate of Mycobacterium tuberculosis

The purification of immunodominant native protein antigens from the culture filtrates of Mycobacterium tuberculosis is needed for the development of new vaccines and immunodiagnostic reagents against tuberculosis. In the present study, we conducted large scale purification of well-known secreted antigens, Ag85 complex, 38-kDa, and MTB12, from the culture filtrate proteins (CFPs) prepared from M. tuberculosis H37Rv grown as a surface pellicle on synthetic Sauton medium. The protein and antigen concentrations of culture filtrates were sufficiently increased after 6 week of culture. The MTB12 antigen was detected as early as 1 week of culture, and Ag85 complex and 38-kDa antigen were detected after 2 and 3 week of culture, respectively, by immunodiffusion with specific antiserum against 100-fold concentrated culture filtrates. For large-scale purification, the six-week-culture filtrates of M. tuberculosis H37Rv diluted 2.5-fold with 20 mM Tris-HCl, (P)H 8.3 were subjected to anion-exchange chromatography. The CFPs were eluted with 100 mM NaCl-20 mM Tris-HCl, pH 8.3 and concentrated by ultrafiltration. The concentrated CFPs were fractionated with ammonium sulfate, and followed by hydrophobic interaction chromatography and anion-exchange chromatography (FPLC). Eventually, 10 mg of Ag85 complex, 0.56 mg of 38-kDa, and 1.81 mg of MTB12 antigens were purified from 1 liter of the six-week-culture filtrates of M. tuberculosis H37Rv which contained 307.81 mg of protein of culture filtrate.

Identification of Proteins Induced at Hypoxic and Low pH Conditions in Mycobacterium tuberculosis H37Rv

Mycobacterium tuberculosis likely reside within a granuloma as a dormant state. An area of necrosis forms at the center of lung granulomas. Within this area, the bacteria are deprived of nutrients and exposed to harsh conditions, including low pH and anoxia. The response of M. tuberculosis to low pH and low oxygen conditions was investigated in both cellular and extracellular proteins by two-dimensional polyacrylamide gel electrophoresis analysis and MALDITOF. Several proteins intensively expressed under low pH and/or hypoxic conditions were found. In the culture filtrate, PhoS1 (Rv0934) and ScoB (Rv2503c) were found in significant amounts under both the low oxygen and acidic stress conditions. These results indeed extend our understanding of acidic response as well as hypoxic in M. tuberculosis and provide an important insight into physiology of the latent bacilli.

The Phospholipase-Protein Kinase C-MEK-ERK Pathway is Essential in Mycobacteria-induced CCL3 and CCL4 Expression in Human Monocytes

BACKGROUND: Little information is available on the identification and characterization of the upstream regulators of the signal transduction cascades for Mycobacterium tuberculosis (M. tbc)-induced ERK 1/2 activation and chemokine expression. We investigated the signaling mechanisms involved in expression of CCL3/MIP-1 and CCL4/MIP-1 in human primary monocytes infected with M. tbc. METHODS: MAP kinase phosphorylation was determined using western blot analysis with specific primary antibodies (ERK 1/2, and phospho-ERK1/2), and the upstream signaling pathways were further investigated using specific inhibitors. RESULTS: An avirulent strain, M. tbc H37Ra, induced greater and more sustained ERK 1/2 phosphorylation, and higher CCL3 and CCL4 production, than did M. tbc H37Rv. Specific inhibitors for mitogen-activated protein kinase (MAPK) kinase (MEK; U0126 and PD98059) significantly inhibited the expression of CCL3 and CCL4 in human monocytes. Mycobacteria-mediated expression of CCL3 and CCL4 was not inhibited by the Ras inhibitor manumycin A or the Raf-1 inhibitor GW 5074. On the other hand, phospholipase C (PLC) inhibitor (U73122) and protein kinase C (PKC)- specific inhibitors (GO6976 and Ro31-8220) significantly reduced M. tbc-induced activation of ERK 1/2 and chemokine synthesis. CONCLUSION: These results are the first to demonstrate that the PLC-PKC-MEK-ERK, not the Ras-Raf-MEK-ERK, pathway is the major signaling pathway inducing M. tbc-mediated CCL3 and CCL4 expression in human primary monocytes.

Induction of TNF-alpha, IL-12, IL-10, MCP-1 and IL-8 in Human Peripheral Blood Monocytes in Response to the Cell Wall-Associated Triton X-100 Soluble Protein (TSP) Antigens of Mycobacterium tuberculosis

Mycobacterium tuberculosis is a potent inducer of cytokine production by mononuclear phagocytes, which are an important cellular component in the first line immune defence. In this study, the cell wall-associated Triton X-100 soluble protein (TSP) antigens, TSP-H37Rv, TSP-H37Ra, TSP-K, and TSP-BCG, were isolated from M. tuberculosis H37Rv, M. tuberculosis H37Ra, M. tuberculosis K-strain, and M. bovis BCG, respectively. The monocytes were isolated from the peripheral blood mononuclear cells of healthy individuals and were co-cultured with each TSP antigens and the secretory proteins of M. tuberculosis (PPD and 30-kDa antigen) to measure the production of cytokines; tumor necrosis factor (TNF)-a, interleukin (IL)-12, IL-8 and monocyte chemotactic protein-1 (MCP-1). The TSP-H37Rv antigen- stimulated monocytes showed higher level of TNF-a and IL-12 production compared to those of other TSP antigens and PPD. Especially, IL-12 production in response to the TSP-H37Rv antigen was significantly elevated in comparison with that of PPD-stimulated monocytes (TSP-H37Rv, 255.5+/-256.9 pg/ml; PPD, 55.7+/-55.4 pg/ml). However, the 30-kDa antigen did not induce TNF-alpha expression and also showed the lowest level of cytokine and chemokine production by monocytes. MCP-1 and IL-8 production were similarly increased in response to all TSP antigens and the PPD antigen. The production of IL-12 by the TSP-H37Rv antigen stimulation was significantly increased in PPD reactors than that in the non-reactor group, while the levels of other cytokines stimulated with each TSP antigens, 30-kDa and PPD antigen were not significantly different between the tuberculin reactor and the non-reactor groups. These results suggest that the cell wall-associated TSP antigen isolated from M. tuberculosis H37Rv acts as a more potent IL-12 inducer than the PPD antigen in innate immune response and thus it could further activate the Th1-mediated immune responses effectively against M. tuberculosis infection.

Mycobacteria-induced Interleukin-12 Expression by Human Monocyte-derived Macrophages Is Negatively Regulated by Phosphatidylinositol 3-kinase and ERK 1/2 Pathways

Both interleukin (IL)-12, an important cytokine skewing the immune response towards a Th1 cytokine profiles, and tumor necrosis factor (TNF)-alpha, are thought to be critical factors in defenses against mycobacteria. In this study, we evaluated the roles of phosphatidylinositol 3-kinase (PI 3-K), and extracellular signal-regulated kinase (ERK) 1/2 pathways in the expression of IL-12 in human monocyte-derived macrophages (MDMs) after stimulation with Mycobacterium tuberculosis H37Rv (M. tbc) or the Triton X-114 solublized proteins (TSP) of M. tbc. Both M. tbc and TSP rapidly phosphorylated ERK 1/2, and Akt in human MDMs. Inhibition of PI 3-K-Akt pathway by specific inhibitors (LY294002 and wortmannin) dramatically increased M. tbc- or TSP-induced IL-12 p40 and p35 mRNA and IL-12 production. In addition, blockade of ERK 1/2 pathway by specific inhibitors (PD98059 and U0126) significantly increased the mRNA levels and cytokine production in M. tbc- or TSP-treated MDMs. On the contrary, M. tbc- or TSP-induced TNF-a production was significantly depressed in human MDMs by pretreatment with inhibitors of PI 3-K or ERK pathways. The M. tbc or TSP stimulation decreased ERK 1/2 phosphorylation by 70% in the presence of wortmannin or LY294002, suggesting that some cross-talk between the PI 3-K-Akt and mitogen-activated protein kinase kinase (MEK)-ERK pathways may be operating in human monocytes during mycobacterial infection. PI 3-K activity is partially required for the M. tbc- or TSP-induced ERK 1/2 phosphorylation. Collectively, these data suggest that the PI 3-K and ERK 1/2 pathways play a central role in the negative regulation of IL-12, but not TNF-a, production by M. tbc.